Kyasanur Forest Disease (KFD) is a tick-borne, zoonotic viral hemorrhagic fever, previously known to be endemic to the state of Karnataka, India. The first outbreak of KFD in Goa state was reported ...in the Sattari taluka, in North Goa in 2015. This study aimed to investigate the outbreak and report the clinical manifestations and risk factors in people diagnosed with KFD.
A mixed methods approach was used, which included a case series report and 19 in-depth interviews (IDIs) conducted with people diagnosed with KFD. The recorded IDIs were transcribed and translated and themes were coded for the analysis.
There were 73 suspected cases of which 30 were confirmed to have KFD using RT-PCR. There were four suspected deaths of which two were confirmed by RT-PCR. Most of the affected individuals were found to be dependent on the forest for their livelihood. Most of the people in the region were engaged in cashew plantations and had to travel to the forest to fetch firewood and cashew, hence were at a higher risk. They lived near the forest. The lack of hemorrhagic manifestation was noteworthy in the current outbreak.
The 'One Health' approach should be implemented to control KFD. Tick bite prevention measures coupled with vaccination of high-risk groups and intensive health education should be carried out, especially before the transmission season. There is a need to have high clinical suspicion for KFD in the region bearing in mind the non-hemorrhagic manifestation in this outbreak.
The structural integrity of the host red blood cell (RBC) is crucial for propagation of Plasmodium spp. during the disease-causing blood stage of malaria infection. To assess the stability of ...Plasmodium vivax-infected reticulocytes, we developed a flow cytometry-based assay to measure osmotic stability within characteristically heterogeneous reticulocyte and P. vivax-infected samples. We find that erythroid osmotic stability decreases during erythropoiesis and reticulocyte maturation. Of enucleated RBCs, young reticulocytes which are preferentially infected by P. vivax, are the most osmotically stable. P. vivax infection however decreases reticulocyte stability to levels close to those of RBC disorders that cause hemolytic anemia, and to a significantly greater degree than P. falciparum destabilizes normocytes. Finally, we find that P. vivax new permeability pathways contribute to the decreased osmotic stability of infected-reticulocytes. These results reveal a vulnerability of P. vivax-infected reticulocytes that could be manipulated to allow in vitro culture and develop novel therapeutics.
Abstract
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 × 10
5
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.
The MCI has laid down a basic framework for interns, which it expects all prospective doctors to be well versed in. Asking students to demonstrate their understanding of the subject, ability to think ...critically, analyze, infer and act accordingly is imperative to the learning process.
To assess competency levels in medical interns post internship via a questionnaire developed based on MCI framework and departmental expectations of clinical capabilities.
This descriptive cross-sectional study was done in a tertiary care hospital involving 74 interns nearing end of internship in the year 2017.
A questionnaire consisting of core competencies such as professionalism, communication skills, learning competency, clinical problem solving amongst others was provided to each and competency levels were assessed against a pre-defined scale. Answers were graded as Poor, Average, Good and Excellent with corresponding numerical equivalents 0, 1, 2 and 3 respectively.
The values obtained were analysed using Weighted Sum technique through Microsoft Excel tool.
An ideal average competency score was initially established and overall competency of each intern was adjudged against the same. Out of 74 candidates that answered the questionnaire, a vast majority of 50 were found at below average competency. Cardio pulmonary resuscitation was known only to 13 students. Around 50 students were severely lacking with regards to knowledge about the use of preferred antibiotic in sepsis and seizures.
There seems to exist significant disconnect in the expectations of MCI on one hand and actual knowledge and skill acquisition of the doctors on other. A departmental wise curriculum and exams at the end of each departmental posting which is more skill based will enable a well-trained doctor with reasonable skills and knowledge to obtain his license to practice.
Efforts to study the biology of Plasmodium vivax liver stages, particularly the latent hypnozoites, have been hampered by the limited availability of P. vivax sporozoites. Anopheles stephensi is a ...major urban malaria vector in Goa and elsewhere in South Asia. Using P. vivax patient blood samples, a series of standard membrane-feeding experiments were performed with An. stephensi under the US NIH International Center of Excellence for Malaria Research (ICEMR) for Malaria Evolution in South Asia (MESA). The goal was to understand the dynamics of parasite development in mosquitoes as well as the production of P. vivax sporozoites. To obtain a robust supply of P. vivax sporozoites, mosquito-rearing and mosquito membrane-feeding techniques were optimized, which are described here.
Membrane-feeding experiments were conducted using both wild and laboratory-colonized An. stephensi mosquitoes and patient-derived P. vivax collected at the Goa Medical College and Hospital. Parasite development to midgut oocysts and salivary gland sporozoites was assessed on days 7 and 14 post-feeding, respectively. The optimal conditions for mosquito rearing and feeding were evaluated to produce high-quality mosquitoes and to yield a high sporozoite rate, respectively.
Laboratory-colonized mosquitoes could be starved for a shorter time before successful blood feeding compared with wild-caught mosquitoes. Optimizing the mosquito-rearing methods significantly increased mosquito survival. For mosquito feeding, replacing patient plasma with naïve serum increased sporozoite production > two-fold. With these changes, the sporozoite infection rate was high (> 85%) and resulted in an average of ~ 22,000 sporozoites per mosquito. Some mosquitoes reached up to 73,000 sporozoites. Sporozoite production could not be predicted from gametocyte density but could be predicted by measuring oocyst infection and oocyst load.
Optimized conditions for the production of high-quality P. vivax sporozoite-infected An. stephensi were established at a field site in South West India. This report describes techniques for producing a ready resource of P. vivax sporozoites. The improved protocols can help in future research on the biology of P. vivax liver stages, including hypnozoites, in India, as well as the development of anti-relapse interventions for vivax malaria.
As much as 80% of global Plasmodium vivax infections occur in South Asia and there is a shortage of direct studies on infectivity of P. vivax in Anopheles stephensi, the most common urban mosquito ...carrying human malaria. In this quest, the possible effects of laboratory colonization of mosquitoes on infectivity and development of P. vivax is of interest given that colonized mosquitoes can be genetically less divergent than the field population from which they originated.
Patient-derived P. vivax infected blood was fed to age-matched wild and colonized An. stephensi. Such a comparison requires coordinated availability of same-age wild and colonized mosquito populations. Here, P. vivax infection are studied in colonized An. stephensi in their 66th-86th generation and fresh field-caught An. stephensi. Wild mosquitoes were caught as larvae and pupae and allowed to develop into adult mosquitoes in the insectary. Parasite development to oocyst and sporozoite stages were assessed on days 7/8 and 12/13, respectively.
While there were batch to batch variations in infectivity of individual patient-derived P. vivax samples, both wild and colonized An. stephensi were roughly equally susceptible to oocyst stage Plasmodium infection. At the level of sporozoite development, significantly more mosquitoes with sporozoite load of 4+ were seen in wild than in colonized populations.
Diabetes is a major public health problem in our country and complications of diabetes are a major cause of morbidity and mortality. There is a need to quantify the complications in order to improve ...our strategies for prevention and management.
To measure the prevalence of complications in type 2 diabetics following up at a tertiary care centre and to study its association with the socio-demographic and clinical parameters.
A retrospective record based study was conducted on 3261 type 2 diabetic patients on insulin therapy, recorded in the diabetic registry maintained at Goa Medical College from Aug 2009 to May 2012. Data on anthropometric measurements, demographic characteristics, complications and other details were extracted from these records.
Out of the 3261 patients 1025 (31.4%) had macrovascular complications and 1122 (34.4%) had at least one microvascular complication. The prevalence of peripheral vascular disease, coronary artery disease and stroke were 6.7%, 21.3% and 6.6% respectively and were significantly higher in males. The prevalence of diabetic retinopathy, nephropathy and neuropathy were 16.7%, 16.5% and 16.3% respectively with diabetic nephropathy being significantly higher in males. Trend analysis showed significant association of rising prevalence of all complications with age (p<0.05). Duration of diabetes also showed significantly positive trend for all complications (p<0.05) except stroke.
The study presents the prevalence of diabetic complications in patients reporting to a tertiary hospital in Goa. Coronary artery disease was found to be the most common complication. As age and duration of diabetes were found to be significantly associated, efforts should be made towards promoting earlier diagnosis of diabetes so as to improve management and decrease the chances of complications.
In global efforts to track mosquito infectivity and parasite elimination, controlled mosquito-feeding experiments can help in understanding the dynamics of parasite development in vectors. Anopheles ...stephensi is often accepted as the major urban malaria vector that transmits Plasmodium in Goa and elsewhere in South Asia. However, much needs to be learned about the interactions of Plasmodium vivax with An. stephensi. As a component of the US NIH International Center of Excellence for Malaria Research (ICEMR) for Malaria Evolution in South Asia (MESA), a series of membrane-feeding experiments with wild An. stephensi and P. vivax were carried out to better understand this vector-parasite interaction.
Wild An. stephensi larvae and pupae were collected from curing water in construction sites in the city of Ponda, Goa, India. The larvae and pupae were reared at the MESA ICEMR insectary within the National Institute of Malaria Research (NIMR) field unit in Goa until they emerged into adult mosquitoes. Blood for membrane-feeding experiments was obtained from malaria patients at the local Goa Medical College and Hospital who volunteered for the study. Parasites were counted by Miller reticule technique and correlation between gametocytaemia/parasitaemia and successful mosquito infection was studied.
A weak but significant correlation was found between patient blood gametocytaemia/parasitaemia and mosquito oocyst load. No correlation was observed between gametocytaemia/parasitaemia and oocyst infection rates, and between gametocyte sex ratio and oocyst load. When it came to development of the parasite in the mosquito, a strong positive correlation was observed between oocyst midgut levels and sporozoite infection rates, and between oocyst levels and salivary gland sporozoite loads. Kinetic studies showed that sporozoites appeared in the salivary gland as early as day 7, post-infection.
This is the first study in India to carry out membrane-feeding experiments with wild An. stephensi and P. vivax. A wide range of mosquito infection loads and infection rates were observed, pointing to a strong interplay between parasite, vector and human factors. Most of the present observations are in agreement with feeding experiments conducted with P. vivax elsewhere in the world.
Parkinson's disease (PD) is a neurodegenerative disorder characterised by the degeneration of dopaminergic nigrostriatal pathway. Its incidence ranges from 1 to 2 per 1000 in general population. Its ...a complex disorder characterised by motor and non-motor symptoms that can impair the quality of life (QoL). In the presence of effective symptomatic therapies for the motor symptoms of Parkinson's disease, the non-motor symptoms have become the major prognostic factors determining the overall disease burden and everyday function in Parkinson’s disease patients.
Our study aimed at assessing the frequency and proportion of non-motor symptoms in PD patients and to assess their impact on QoL.
A total of 88 PD patients visiting the outpatient department of Neurology at Goa Medical College were assessed for various non-motor symptoms using a semi – structured questionnaire and appropriate scales. Quality of life was assessed using PDQ-39 questionnaire. Patients satisfying the United Kingdom Parkinson’s Disease brain bank criteria for idiopathic Parkinson’s diseases were included in the study
Single centre, Cross-sectional study.
Mean age of study population was 68 years and mean duration of the disease was 5yrs. Attention and memory were the most frequently reported domain in the study population (52 patients, 59.75 percent) followed by mood and cognition (46 patients, 52.74 percent). The least reported symptom was perceptual problems including delusions and hallucinations (11.74%) and miscellaneous domain (10.93 percent) which included pain, olfactory dysfunction, increased perspiration and weight fluctuations. The mean UPDRS (part 3) score was found to be 33.24± 17.37 with values ranging from 3.57 (mild motor impairment) to 44.64 (severe motor impairment).
The QoL assessment showed that impairment of Mobility had a major impact on QoL followed by activities of daily living and emotional well-being. Correlation between Non-motor symptoms and PDQ-39 revealed that mood, cognition and perceptual problems had an adverse outcome on their emotional well-being and deficit in attention and memory had significant outcomes on communication.
This study highlights the fact that non-motor symptoms in PD significantly impair quality of life and we should incorporate screening tools to evaluate patients with autonomic, sensory or mood disturbances so that they are detected at the earliest as most of them precede motor symptoms by decades.
Chikungunya is an arboviral disease transmitted by Aedes mosquito that represents a major public health burden worldwide including India. The disease presents as sudden onset of high-grade fever, ...severe arthralgias, and rash. Here, we describe a case of a patient who presented with cervical lymphadenopathy, fever, and myalgia and later was diagnosed as chikungunya. Lymphadenopathy has been described before as a less common symptom of chikungunya. But this is probably, the first case of chikungunya with cervical lymphadenopathy as a presenting feature.
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