Correspondence: The African snakebite Alliance.

The goal to reduce the burden of snakebite envenoming is challenged by the gaps in evidence for clinical care and public health. These evidence gaps and the absence of a strong network are illustrated by bibliometrics. The African Snakebite Alliance is a multidisciplinary group focusing on research themes which will generate evidence needed to shape policy and practice.

Stakeholder perspectives from 15 countries in Africa on barriers in snakebite envenoming research and the potential role of research hubs.

Snakebite envenoming is a debilitating neglected tropical disease disproportionately affecting the rural poor in low and middle-income countries in the tropics and sub-tropics. Critical questions and gaps in public health and policy need to be addressed if major progress is to be made towards reducing the negative impact of snakebite, particularly in the World Health Organisation (WHO) Africa region. We engaged key stakeholders to identify barriers to evidence-based snakebite decision making and to explore how development of research and policy hubs could help to overcome these barriers. We conducted an electronic survey among 73 stakeholders from ministries of health, health facilities, academia and non-governmental organizations from 15 countries in the WHO Africa region. The primary barriers to snakebite research and subsequent policy translation were limited funds, lack of relevant data, and lack of interest from policy makers. Adequate funding commitment, strong political will, building expert networks and a demand for scientific evidence were all considered potential factors that could facilitate snakebite research. Participants rated availability of antivenoms, research skills training and disease surveillance as key research priorities. All participants indicated interest in the development of research and policy hubs and 78% indicated their organization would be willing to actively participate. In conclusion, our survey affirms that relevant stakeholders in the field of snakebite perceive research and policy hubs as a promising development, which could help overcome the barriers to pursuing the WHO goals and targets for reducing the burden of snakebite.

Snakebite epidemiology, outcomes and multi-cluster risk modelling in Eswatini.

BACKGROUND: Halving snakebite morbidity and mortality by 2030 requires countries to develop both prevention and treatment strategies. The paucity of data on the global incidence and severity of snakebite envenoming causes challenges in prioritizing and mobilising resources for snakebite prevention and treatment. In line with the World Health Organisation's 2019 Snakebite Strategy, this study sought to investigate Eswatini's snakebite epidemiology and outcomes, and identify the socio-geographical factors associated with snakebite risk. METHODOLOGY: Programmatic data from the Ministry of Health, Government of Eswatini 2019-2021, was used to assess the epidemiology and outcomes of snakebite in Eswatini. We developed a snake species richness map from the occurrence data of all venomous snakes of medical importance in Eswatini that was subjected to niche modelling. We formulated four risk indices using snake species richness, various geospatial datasets and reported snakebites. A multivariate cluster modelling approach using these indices was developed to estimate risk of snakebite and the outcomes of snakebite in Eswatini. PRINCIPAL FINDINGS: An average of 466 snakebites was recorded annually in Eswatini. Bites were recorded across the entire country and peaked in the evening during summer months. Two cluster risk maps indicated areas of the country with a high probability of snakebite and a high probability of poor snakebite outcomes. The areas with the highest rate of snakebite risk were primarily in the rural and agricultural regions of the country. SIGNIFICANCE: These models can be used to inform better snakebite prevention and treatment measures to enable Eswatini to meet the global goal of reducing snakebite morbidity and mortality by 50% by 2030. The supply chain challenges of antivenom affecting southern Africa and the high rates of snakebite identified in our study highlight the need for improved snakebite prevention and treatment tools that can be employed by health care workers stationed at rural, community clinics.

Two snakebite antivenoms have potential to reduce Eswatini's dependency upon a single, increasingly unavailable product: Results of preclinical efficacy testing.

BACKGROUND: Snakebite is a major public health concern in Eswatini, where treatment relies upon one antivenom-SAIMR Polyvalent. Although effective in treating snakebite, SAIMR Polyvalent is difficult to source outside its manufacturing country (South Africa) and is dauntingly expensive. We compared the preclinical venom-neutralising efficacy of two alternative antivenoms with that of SAIMR Polyvalent against the lethal and tissue-destructive effects of venoms from five species of medically important snakes using in vivo murine assays. The test antivenoms were 'Panafrican' manufactured by Instituto Clodomiro Picado and 'PANAF' manufactured by Premium Serums & Vaccines. PRINCIPAL FINDINGS: In vivo murine preclinical studies identified both test antivenoms were equally or more effective than SAIMR Polyvalent at neutralising lethal and tissue-destructive effects of Naja mossambica venom. Both test antivenoms were less effective than SAIMR Polyvalent at neutralising the lethal effects of Bitis arietans, Dendroaspis polylepis, Hemachatus haemachatus and Naja annulifera venoms, but similarly effective at neutralising tissue damage induced by B. arietans and H. haemachatus venoms. In vitro immunological assays identified that the titres and toxin-specificities of immunoglobulins (iGs) in the test antivenoms were comparable to that of SAIMR Polyvalent. Plasma clotting disturbances by H. haemachatus and N. mossambica were neutralised by the test antivenoms, whereas SAIMR Polyvalent failed to neutralise this bioactivity of N. mossambica venom. B. arietans SVMP activity was equally reduced by all three antivenoms, and H. haemachatus and N. mossambica PLA2 activities were neutralised by all three antivenoms. CONCLUSIONS: While both Panafrican and PANAF antivenoms exhibited promising preclinical efficacies, both were less poly-specifically effective than SAIMR Polyvalent in these murine assays. The efficacy of these antivenoms against the lethal and tissue-destructive effects of N. mossambica venom, the most common biting species in Eswatini, identify that Panafrican and PANAF antivenoms offer effective alternatives to SAIMR Polyvalent for the treatment of snakebite in Eswatini, and potentially for neighbouring countries.

Assessment of early COVID-19 compliance to and challenges with public health and social prevention measures in the Kingdom of Eswatini, using an online survey.

Public health and social measures have been implemented around the world in a bid to prevent the spread of COVID-19. Public compliance with these measures is key in successfully controlling the pandemic. This online survey assessed the compliance and attitude of adults residing in the southern African Kingdom of Eswatini to government protection, activity and travel measures aimed at controlling the spread of COVID-19. A rapid online survey, comprising of 28 questions, was administered in May 2020. More than 90% of respondents knew the virus could kill anyone and most respondents (70%) reported to be compliant to public health and social measures. Females, those who did not use public transport and those aged 30 years and above were significantly (p<0.01) more compliant, particularly to protective and travel measures. Social media, television and official government websites were the primary source of ongoing COVID-19 information for respondents of this online survey, and these methods should continue to be employed to reach the public who regularly use the internet. More than half of essential workers who responded to the online survey reported to have their own personal protective equipment; however, 32% either did not have any protective equipment or shared their equipment with other staff members. Due to the survey being online, these results should not be generalised to populations of low socioeconomic status.

High-fat diet alters gene expression in the liver and colon: links to increased development of aberrant crypt foci.

BACKGROUND: Obesity is associated with an increased risk of colon cancer. High-fat diets that lead to obesity may be a contributing factor, but the mechanisms are unknown. AIMS: This study examines susceptibility to azoxymethane (AOM)-induced precancerous lesions in mice in response to consumption of either a low or a high-fat diet and associated molecular changes in the liver and colon. METHODS: Gene markers of xenobiotic metabolism, leptin-regulated inflammatory cytokines and proliferation were assessed in liver and colon in response to high-fat feeding to determine links with increased sensitivity to AOM. RESULTS: High-fat feeding increased development of AOM-induced precancerous lesions and was associated with increased CYP2E1 gene expression in the liver, but not the colon. Leptin receptors and the colon stem cell marker (Lgr5) were down-regulated in the proximal colon, with a corresponding up-regulation of the inflammatory cytokine (IL6) in response to high-fat feeding. Notably in the distal colon, where aberrant crypt foci develop in response to AOM, the proliferative stem cell marker, Lgr5, was significantly up-regulated with high-fat feeding. CONCLUSIONS: The current study provides evidence that high-fat diets can alter regulation of molecular markers of xenobiotic metabolism that may expose the colon to carcinogens, in parallel with activation of ß-catenin-regulated targets regulating colon epithelial cells. High-fat diets associated with obesity may alter multiple molecular factors that act synergistically to increase the risk of colon cancer associated with obesity.

Leptin up-regulates pro-inflammatory cytokines in discrete cells within mouse colon.

Dysregulation of leptin associated with obesity is implicated in obesity-related colon cancer, but mechanisms are elusive. Increased adiposity and elevated plasma leptin are associated with perturbed metabolism in colon and leptin receptors are expressed on colon epithelium. We hypothesise that obesity increases the sensitivity of the colon to cancer by disrupting leptin-regulated gene targets within colon tissues. PCR arrays were used to firstly identify leptin responsive genes and secondly to identify responses to leptin challenge in wild-type mice, or those lacking leptin (ob/ob). Leptin-regulated genes were localised in the colon using in situ hybridisation. IL6, IL1ß and CXCL1 were up-regulated by leptin and localised to discrete cells in gut epithelium, lamina propria, muscularis and at the peritoneal serosal surface. Leptin regulates pro-inflammatory genes such as IL6, IL1ß and CXCL1, and might increase the risk of colon cancer among obese individuals.

Impact of obesity and leptin on protein expression profiles in mouse colon.

BACKGROUND: Elevated leptin levels in obesity are associated with increased risk of colon pathology, implicating leptin signaling in colon disease. However, leptin-regulated processes in the colon are currently uncharacterized. Previously, we demonstrated that leptin receptors are expressed on colon epithelium and that increased adiposity and elevated plasma leptin in rats are associated with perturbed metabolism in colon tissue. Thus, we hypothesize that obesity disrupts expression of proteins regulated by leptin in the colon. METHODS: A proteomic analysis was conducted to investigate firstly, differences in the colon of mice lacking leptin and leptin signaling (ob/ob and db/db, respectively) by comparing protein expression profiles with wild-type mice. Secondly, responses to leptin challenge in wild-type mice and ob/ob mice were compared to identify leptin-regulated proteins and associated cellular processes. RESULTS: Forty proteins were identified with significantly altered expression patterns associated with differences in leptin status in comparisons between all groups of mice. These proteins are associated with calcium binding, cell cycle, cell proliferation, electron transport chain, energy metabolism, protein folding and transport, redox regulation, structural proteins, and proteins involved in transport and regulation of mucus production. CONCLUSIONS: This study provides evidence that obesity and leptin significantly alter protein profiles of a number of proteins linked to cellular processes in colon tissues that may be linked to the increased risk of colon pathology associated with obesity.

Influence of increased adiposity on mitochondrial-associated proteins of the rat colon: a proteomic and transcriptomic analysis.

Epidemiological studies report obesity to be an important risk factor influencing colon pathologies, yet mechanism(s) are unknown. Recent studies have shown significant elevation of insulin, leptin and triglycerides associated with increased adipose tissue. In situ hybridisation studies have located insulin, leptin and adiponectin receptor expression in the colon epithelia. The influence of increased adiposity and associated deregulation of insulin and adipokines on regulation of the colon epithelium is unknown. Altered adipokine and insulin signalling associated with obesity has an impact on mitochondrial function and mitochondrial dysfunction is increasingly recognised as a contributing factor in many diseases. Proteomics and transcriptomics are potentially powerful methods useful in elucidating the mechanisms whereby obesity increases risk of colon diseases as observed epidemiologically. This study investigated colon mitochondrial-associated protein profiles and corresponding gene expression in colon in response to increased adiposity in a rat model of diet induced obesity. Increased adiposity in diet-induced obese sensitive rats was found to be associated with altered protein expression of 69 mitochondrial-associated proteins involved in processes associated with calcium binding, protein folding, energy metabolism, electron transport chain, structural proteins, protein synthesis and degradation, redox regulation, and transport. The changes in these mitochondrial protein profiles were not correlated with changes at the gene expression level assessed using real-time PCR arrays.

Profiling of mitochondrial associated proteins from rat colon.

Mitochondrial dysfunction, damage and mutations of mitochondrial proteins give rise to a range of ill understood patterns of disease. Although there is significant general knowledge of the proteins and the functional processes of the mitochondria, there is little knowledge of difference about how mitochondria respond and how they are regulated in different organs and tissues. Proteomic profiling of mitochondria and associated proteins involved in mitochondrial regulation and trafficking within cells and tissues has the potential to provide insights into mitochondrial dysfunction associated with many human diseases. The rat colon mitoproteome analysis presented here provides a useful tool to assist in identification and interpretation of mitochondrial dysfunction implicated in colon pathogenesis. 2DPAGE followed by LC/MS/MS was used to identify 430 proteins from mitochondrial enriched fractions prepared from rat colon, resulting in 195 different proteins or approximately 50% of the resolved proteins being identified as multiple protein expression forms. Proteins associated with the colon mitoproteome were involved in calcium binding, cell cycle, energy metabolism and electron transport chain, protein folding, protein synthesis and degradation, redox regulation, structural proteins, signalling and transporter and channel proteins. The mitochondrial associated proteins identified in this study of colon tissue complement and are compared with other recently published mitoproteome analyses from other organ tissues, and will assist in revealing potentially organ specific roles of the mitochondria and organ specific disease associated with mitochondrial dysfunction.

Insulin, leptin, and adiponectin receptors in colon: regulation relative to differing body adiposity independent of diet and in response to dimethylhydrazine.

Obesity has recently become a focus of research to elucidate diet and lifestyle factors as important risk factors for colon cancer. Altered levels of insulin, leptin, and adiponectin have been identified as potential candidates increasing colon cancer risk within the prevailing obesogenic environment. There has been considerable research to characterize signaling via these hormones in the brain, liver, and adipose tissue; however, very little is known of their emerging role in peripheral signaling, particularly in epithelial tissues. This study profiles insulin, leptin, and adipokine receptors in the rat colon, revealing novel microanatomical location of these receptors and thereby supporting a potential role in regulating colonic tissue. Potential involvement of insulin, leptin, and adiponectin receptors in increased risk of colon cancer was investigated using Sprague-Dawley rats, either resistant or susceptible to diet-induced obesity. Regulation of insulin, leptin, and adiponectin receptors as a consequence of differing levels of adiposity was assessed regionally in the colon in response to treatment with the chemical carcinogen 1,2-dimethylhydrazine (DMH). However, significantly increased fat mass, increased levels of plasma insulin, leptin, and triglycerides, previously associated with an increased risk of colon cancer, were not associated with promotion of precancerous lesions in the experimental rats or deregulation of insulin, leptin, or adiponectin receptors. These findings do not support a direct link between the deregulation of insulin and adipokine levels observed in obese rats and an increased risk of colon carcinogenesis.

Salicylate modulates oxidative stress in the rat colon: a proteomic approach.

The dietary phenolic compound, salicylic acid, decreases oxidative stress and pro-inflammatory and potentially neo-plastic prostaglandins with a concomitant increase in glutathione peroxidase activity. Salicylic acid, a dietary plant-based phenolic compound and also the main metabolite of aspirin, may contribute to the colon protective effects of plant-based diets. Oxidative stress is a characteristic of pre-cancerous and cancerous colon and inflammatory bowel diseases (IBD) that increase colon cancer risk. The mechanism(s) whereby salicylic acid modulates potentially pro-cancerous activity associated with oxidative stress is further investigated here using a proteomic approach. A rat model of oxidative stress was supplemented with salicylic acid (1 mg/kg diet, mean plasma levels 310+/-32 micromol/l). Soluble colon protein extracts were subjected to 2D PAGE. Salicylic acid modulated proteins, identified using MALDI-TOF and LC/MS/MS, are involved in protein folding, transport, redox, energy metabolism and cytoskeletal regulation. A partial least squares (PLS) analysis approach was used to assist biological interpretation of the altered protein profiles via their associations between previously published biochemical measurements of oxidative stress, prostaglandin levels and increased glutathione peroxidase activity. Early detection of altered homeostasis in colon may assist in identifying pre-pathological features preceding colon tumorigenesis and contribute to an understanding of epidemiological evidence supporting a protective effect of plant-based diets.