RESUMO
BACKGROUND: Plasmodium vivax is the second most prevalent cause of malaria yet remains challenging to study due to the lack of a continuous in vitro culture system, highlighting the need to establish a biobank of clinical isolates with multiple freezes per sample for use in functional assays. Different methods for cryopreserving parasite isolates were compared and subsequently the most promising one was validated. Enrichment of early- and late-stage parasites and parasite maturation were quantified to facilitate assay planning. METHODS: In order to compare cryopreservation protocols, nine clinical P. vivax isolates were frozen with four glycerolyte-based mixtures. Parasite recovery post thaw, post KCl-Percoll enrichment and in short-term in vitro culture was measured via slide microscopy. Enrichment of late-stage parasites by magnetic activated cell sorting (MACS) was measured. Short and long-term storage of parasites at either - 80 °C or liquid nitrogen were also compared. RESULTS: Of the four cryopreservation mixtures, one mixture (glycerolyte:serum:RBC at a 2.5:1.5:1 ratio) resulted in improved parasite recovery and statistically significant (P < 0.05) enhancement in parasite survival in short-term in vitro culture. A parasite biobank was subsequently generated using this protocol resulting in a collection of 106 clinical isolates, each with 8 vials. The quality of the biobank was validated by measuring several factors from 47 thaws: the average reduction in parasitaemia post-thaw (25.3%); the average fold enrichment post KCl-Percoll (6.65-fold); and the average percent recovery of parasites (22.0%, measured from 30 isolates). During short-term in vitro culture, robust maturation of ring stage parasites to later stages (> 20% trophozoites, schizonts and gametocytes) was observed in 60.0% of isolates by 48 h. Enrichment of mature parasite stages via MACS showed good reproducibility, with an average of 30.0% post-MACS parasitaemia and an average of 5.30 × 105 parasites/vial. Finally, the effect of storage temperature was tested, and no large impacts from short-term (7 days) or long-term (7-10 years) storage at - 80 °C on parasite recovery, enrichment or viability was observed. CONCLUSIONS: Here, an optimized freezing method for P. vivax clinical isolates is demonstrated as a template for the generation and validation of a parasite biobank for use in functional assays.
Assuntos
Malária Vivax , Plasmodium vivax , Humanos , Bancos de Espécimes Biológicos , Reprodutibilidade dos Testes , ParasitemiaRESUMO
BACKGROUND: Multisystem inflammatory syndrome in adults (MIS-A) is an emergent heterogenous clinical syndrome seen in the convalescent phase of COVID-19 infection. MIS in children (MIS-C) is a rare but severe post-COVID-19 illness that has been recognized by the WHO and the Centre for Disease Control and Prevention (CDC). It introduced a similar illness in adults based on multiple case series, identified as MIS-A. OBJECTIVE: We present four rare cases of multiorgan inflammatory syndrome in adults (MI-A) presented in Goa Medical College (Tertiary Medical Institute). We would like to highlight the diversity of presentation of symptoms with a significant history of previous covid infection, laboratory abnormalities, the clinical course of the disease, treatment strategies, and response and follow-up findings. We seek to highlight the emergence of a serious clinical entity that can be fatal if not diagnosed or treated promptly. MATERIALS AND METHODS: This was a descriptive study conducted in Goa Medical College from June 2021 to November 2021. A systematic search in the Department of General Medicine, the Department of Medical Records, and data from ICU, ITU, and critical covid wards were collected. RESULTS AND CONCLUSION: A total of four cases fulfilling the criteria for MIS-A as per MMWR (CDC 2020)were included, ranging from the age group of 29-70 years. All had features of severe systemic inflammatory response with multiple organ dysfunction and elevated proinflammatory markers. All four patients had a recent history of (mild) COVID-19 infection. Hence, in the current pandemic scenario, MIS-A should be considered as a possible diagnosis in patients with recent COVID infection presenting with MODS, when the obvious septic cause is excluded through thorough clinical, physical, serological, laboratory, and radiological investigations. However, the presence of a past covid infection may not be an absolute criterion due to mild symptoms of the primary covid infection which usually go unnoticed resulting in nontesting.
Assuntos
COVID-19 , Doenças do Tecido Conjuntivo , Adulto , Idoso , Criança , Humanos , Pessoa de Meia-Idade , SARS-CoV-2 , Síndrome , Síndrome de Resposta Inflamatória Sistêmica/diagnóstico , Síndrome de Resposta Inflamatória Sistêmica/terapiaRESUMO
BACKGROUND AND OBJECTIVES: Henoch Schonlein purpura (HSP) is a small vessel vasculitic disorder common in children and has been extensively studied. Although it is known to also occur in adults there is relative paucity of data as regards to its clinical spectrum, complications and outcome, particularly in Indian context. Hence the study was undertaken with the objective to evaluate the various skin manifestations, systemic complications of HSP in adults and also compare it with data available in children in various published clinical studies.Study design, materials, methods : In this retrospective, observational, hospital-based cohort study conducted at Goa Medical College the premier teaching institute from Goa, clinical data of adult patients (>18 years age) fulfilling the diagnostic criteria as per European League Against Rheumatism (EULAR) 2010 criteria for HSP was obtained, over period of 6 years. All the clinical manifestations, complications, investigations, outcomes were recorded. Skin biopsy histopathology and immunofluorescent test findings were also obtained. The data was analysed and results were compared to the data available in pediatric studies to ascertain the similarities and differences. RESULTS: In our study cohort of 30 patients, we found a higher incidence of atypical and more extensive skin lesions particularly bullae (40%), necrotic ulcers (53.3%), urticarial wheals (53%) unlike in children as well as differences in distribution especially sparing of buttocks in adults. The incidence of gastrointestinal involvement was 80% which was higher than that reported by other studies in adults (35% to 70%). A significant 40% of patients had upper GI bleeding with endoscopy revealing small hemorrhages in gastric mucosa. Lower GIT bleed was seen in 8 patients. Renal involvement (microscopic hematuria, overt glomerulonephritis, nephrotic syndrome) was seen in 65% patients which was higher than that reported in children (43 %). Skin biopsy immunofluorescence was found to be positive in almost 66 % cases confirming IgA deposition which is the hallmark pathological finding. CONCLUSIONS: HSP, though less common in adults than children, presents with atypical and more severe cutaneous manifestations like bullae, necrotic ulcers, urticarial wheals. Systemic involvement appears to be more frequent and causing more morbidity and mortality as compared to the data in children mentioned in standard literature and most of the patients required steroid therapy for treatment unlike in children where majority of these cases are self-limiting. Skin involvement does not necessarily mirror gastrointestinal involvement in terms of severity and temporal occurrence.
Assuntos
Vasculite por IgA , Adulto , Vesícula , Criança , Estudos de Coortes , Hospitais , Humanos , Vasculite por IgA/complicações , Vasculite por IgA/diagnóstico , Estudos Retrospectivos , ÚlceraRESUMO
BACKGROUND: Polycythemia is characterized by rise in hemoglobin and hematocrit, either as a result of hematopoietic clonal expansion (Vera) or secondary to hypoxic stimuli (secondary polycythemia).It is of great importance to detect early and identify the type of polycythemia and also asses the thrombotic risk so that timely and appropriate treatment can be given. The present study aims to characterize the different presentations and complications of polycythemia, evaluate genetic factors and differences between the two categories of polycythemia in ethnic Goan subjects. AIMS AND OBJECTIVES: 1. To identify common presentations and etiologies of polycythemia 2. To evaluate and compare the differences in clinical features, hematological parameters and complications of polycythemia in primary (vera) and secondary polycythemia 3. To study the profile of JAK 2 V617F mutation in Goan patients with polycythemia Vera. MATERIALS AND METHODS: This was a retrospective observational cohort study, conducted at the Department of Internal Medicine, Goa Medical College, a tertiary care, teaching institute in the state of Goa. We analysed clinical and laboratory data of patients of polycythemia due to all causes (polycythemia Vera and secondary causes) previously admitted or following up at the hospital from January 2014 to December 2017. In each of these 2 groups, we studied the various clinical parameters including the age at presentation,sex,residence, symptomatology and clinical findings,presence of hypertension, as well as complications arising due to polycythemia (past and at present) hematological data including Hb,HCT, total WBC count, absolute neutrophil count, RBC and platelet count, ESR, rouleaux formation, EPO levels and JAK 2 V617F mutation analysis (done by real time PCR technique) and requirement of phlebotomies in the last 4 years. Commonest clinical presentations and complications arising due to polycythemia, in each group were analysed and compared. RESULTS: A total of 44 patients were included in the study out of which 33 were males. Polycythemia Vera was seen in 43.18% while secondary causes were seen in 56.8 % patients. Patients with Vera were found to be more symptomatic with higher levels of mean Hb, HCT, cell counts and with a higher requirement of phlebotomy and more thrombotic complications. Amongst Vera group, patients having high WBC count, increased Rouleau formation, and JAK2 positivity were found to be more prone for thrombosis. Hypertension was frequently seen to be associated with both groups. Obstructive sleep apnea followed by COPD was found to be the commonest causes of secondary polycythemia. CONCLUSION: Our study revealed that patients with polycythemia Vera are more symptomatic and have a higher requirement of phlebotomy and a higher thrombotic tendency (arterial being more common than venous) as compared to the secondary polycythemia owing to a higher hyperviscocity in the former. Leukocytosis and JAK 2 617F positivity were found to be important predictors of thrombotic risk. Hypertension was found to be frequently associated with Vera as well as in secondary causes due to OSA.
Assuntos
Policitemia/epidemiologia , Estudos de Coortes , Humanos , Índia , Masculino , Policitemia Vera , Estudos Retrospectivos , Centros de Atenção TerciáriaRESUMO
BACKGROUND: Malaria remains an important cause of morbidity and mortality in India. Though many comprehensive studies have been carried out in Africa and Southeast Asia to characterize and examine determinants of Plasmodium falciparum and Plasmodium vivax malaria pathogenesis, fewer have been conducted in India. METHODS: A prospective study of malaria-positive individuals was conducted at Goa Medical College and Hospital (GMC) from 2012 to 2015 to identify demographic, diagnostic and clinical indicators associated with P. falciparum and P. vivax infection on univariate analysis. RESULTS: Between 2012 and 2015, 74,571 febrile individuals, 6287 (8.4%) of whom were malaria positive, presented to GMC. The total number of malaria cases at GMC increased more than two-fold over four years, with both P. vivax and P. falciparum cases present year-round. Some 1116 malaria-positive individuals (mean age = 27, 91% male), 88.2% of whom were born outside of Goa and 51% of whom were construction workers, were enroled in the study. Of 1088 confirmed malaria-positive patients, 77.0% had P. vivax, 21.0% had P. falciparum and 2.0% had mixed malaria. Patients over 40 years of age and with P. falciparum infection were significantly (p < 0.001) more likely to be hospitalised than younger and P. vivax patients, respectively. While approximately equal percentages of hospitalised P. falciparum (76.6%) and P. vivax (78.9%) cases presented with at least one WHO severity indicator, a greater percentage of P. falciparum inpatients presented with at least two (43.9%, p < 0.05) and at least three (29.9%, p < 0.01) severity features. There were six deaths among the 182 hospitalised malaria positive patients, all of whom had P. falciparum. CONCLUSION: During the four year study period at GMC, the number of malaria cases increased substantially and the greatest burden of severe disease was contributed by P. falciparum.
Assuntos
Malária Falciparum/patologia , Malária Vivax/patologia , Adolescente , Adulto , Idoso , Criança , Pré-Escolar , Demografia , Feminino , Humanos , Incidência , Índia/epidemiologia , Lactente , Malária Falciparum/diagnóstico , Malária Falciparum/epidemiologia , Malária Vivax/diagnóstico , Malária Vivax/epidemiologia , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Centros de Atenção Terciária , Adulto JovemRESUMO
The South Asia International Center of Excellence for Malaria Research, an NIH-funded collaborative program, investigated the epidemiology of malaria in the Indian state of Goa through health facility-based data collected from the Goa Medical College and Hospital (GMC), the state's largest tertiary healthcare facility, between 2012 and 2021. Our study investigated region-specific spatial and temporal patterns of malaria transmission in Goa and the factors driving such patterns. Over the past decade, the number of malaria cases, inpatients, and deaths at the GMC decreased significantly after a peak in 2014-2015. However, the proportion of severe malaria cases increased over the study period. Also, a trend of decreasing average parasitemia and increasing average gametocyte density suggests a shift toward submicroscopic infections and an increase in transmission commitment characteristic of low-transmission regions. Although transmission occurred throughout the year, 75% of the cases occurred between June and December, overlapping with the monsoon (June-October), which featured rainfall above yearly average, minimal diurnal temperature variation, and high relative humidity. Sociodemographic factors also had a significant association with malaria cases, with cases being more frequent in the 15-50-year-old age group, men, construction workers, and people living in urban areas within the GMC catchment region. Our environmental model of malaria transmission projects almost negligible transmission at the beginning of 2025 (annual parasitic index: 0.0095, 95% CI: 0.0075-0.0114) if the current control measures continue undisrupted.
Assuntos
Malária , Humanos , Índia/epidemiologia , Adolescente , Feminino , Adulto , Masculino , Criança , Pessoa de Meia-Idade , Adulto Jovem , Pré-Escolar , Lactente , Malária/transmissão , Malária/epidemiologia , Malária/prevenção & controle , Idoso , Estações do Ano , Hospitais/estatística & dados numéricos , Erradicação de Doenças , Malária Falciparum/epidemiologia , Malária Falciparum/transmissão , Malária Falciparum/prevenção & controleRESUMO
Background: Plasmodium vivax is the second most prevalent cause of malaria yet remains challenging to study due to the lack of a continuous in vitro culture system, highlighting the need to establish a biobank of clinical isolates with multiple freezes per sample for use in functional assays. Different methods for cryopreserving parasite isolates were compared and subsequently the most promising one was validated. Enrichment of early- and late-stage parasites and parasite maturation were quantified to facilitate assay planning. Methods: In order to compare cryopreservation protocols, nine clinical P. vivax isolates were frozen with four glycerolyte-based mixtures. Parasite recovery post thaw, post KCl-Percoll enrichment and in short-term in vitro culture was measured via slide microscopy. Enrichment of late-stage parasites by magnetic activated cell sorting (MACS) was measured. Short and long-term storage of parasites at either -80°C or liquid nitrogen were also compared. Results: Of the four cryopreservation mixtures, one mixture (glycerolyte:serum:RBC at a 2.5:1.5:1 ratio) resulted in improved parasite recovery and statistically significant (P<0.05) enhancement in parasite survival in short-term in vitro culture. A parasite biobank was subsequently generated using this protocol resulting in a collection with 106 clinical isolates, each with 8 vials. The quality of the biobank was validated by measuring several factors from 47 thaws: the average reduction in parasitemia post-thaw (25.3%); the average fold enrichment post KCl-Percoll (6.65-fold); and the average percent recovery of parasites (22.0%, measured from 30 isolates). During short-term in vitro culture, robust maturation of ring stage parasites to later stages (>20% trophozoites, schizonts and gametocytes) was observed in 60.0% of isolates by 48 hours. Enrichment of mature parasite stages via MACS showed good reproducibility, with an average 30.0% post-MACS parasitemia and an average 5.30 × 10 5 parasites/vial. Finally, the effect of storage temperature was tested, and no large impacts from short-term (7 day) or long term (7 - 10 year) storage at -80°C on parasite recovery, enrichment or viability was observed. Conclusions: Here, an optimized freezing method for P. vivax clinical isolates is demonstrated as a template for the generation and validation of a parasite biobank for use in functional assays.
RESUMO
The "Malaria Evolution in South Asia" (MESA) program project is an International Center of Excellence for Malaria Research (ICEMR) sponsored by the US National Institutes of Health. This US-India collaborative program will study the origin of genetic diversity of malaria parasites and their selection on the Indian subcontinent. This knowledge should contribute to a better understanding of unexpected disease outbreaks and unpredictable disease presentations from Plasmodium falciparum and Plasmodium vivax infections. In this first of two reviews, we highlight malaria prevalence in India. In particular, we draw attention to variations in distribution of different human-parasites and different vectors, variation in drug resistance traits, and multiple forms of clinical presentations. Uneven malaria severity in India is often attributed to large discrepancies in health care accessibility as well as human migrations within the country and across neighboring borders. Poor access to health care goes hand in hand with poor reporting from some of the same areas, combining to possibly distort disease prevalence and death from malaria in some parts of India. Corrections are underway in the form of increased resources for disease control, greater engagement of village-level health workers for early diagnosis and treatment, and possibly new public-private partnerships activities accompanying traditional national malaria control programs in the most severely affected areas. A second accompanying review raises the possibility that, beyond uneven health care, evolutionary pressures may alter malaria parasites in ways that contribute to severe disease in India, particularly in the NE corridor of India bordering Myanmar Narayanasamy et al., 2012.