MucoRice-CTB line 19A, a new marker-free transgenic rice-based cholera vaccine produced in an LED-based hydroponic system.

We previously established the selection-marker-free rice-based oral cholera vaccine (MucoRice-CTB) line 51A for human use by Agrobacterium-mediated co-transformation and conducted a double-blind, randomized, placebo-controlled phase I trial in Japan and the United States. Although MucoRice-CTB 51A was acceptably safe and well tolerated by healthy Japanese and U.S. subjects and induced CTB-specific antibodies neutralizing cholera toxin secreted by Vibrio cholerae, we were limited to a 6-g cohort in the U.S. trial because of insufficient production of MucoRice-CTB. Since MucoRice-CTB 51A did not grow in sunlight, we re-examined the previously established marker-free lines and selected MucoRice-CTB line 19A. Southern blot analysis of line 19A showed a single copy of the CTB gene. We resequenced the whole genome and detected the transgene in an intergenic region in chromosome 1. After establishing a master seed bank of MucoRice-CTB line 19A, we established a hydroponic production facility with LED lighting to reduce electricity consumption and to increase production capacity for clinical trials. Shotgun MS/MS proteomics analysis of MucoRice-CTB 19A showed low levels of α-amylase/trypsin inhibitor-like proteins (major rice allergens), which was consistent with the data for line 51A. We also demonstrated that MucoRice-CTB 19A had high oral immunogenicity and induced protective immunity against cholera toxin challenge in mice. These results indicate that MucoRice-CTB 19A is a suitable oral cholera vaccine candidate for Phase I and II clinical trials in humans, including a V. cholerae challenge study.

Lessons learned from contamination with endotoxin originated from the supplement in the cell culture medium.

Introduction: Endotoxin is a typical pyrogen derived from the outer membrane of Gram-negative bacteria. In fabricating cell-based medicinal products, it is necessary to control endotoxin in the process and the products. In the quality control tests of our clinical study, endotoxin concentration in the culture supernatant of autologous oral mucosal epithelial cell sheets exceeded the criterion value. Therefore, endotoxin measurements were conducted to clarify the cause of the endotoxin contamination. Methods: The reagents used to prepare the culture medium, the unused culture medium, and the culture supernatants were diluted with pure water. Endotoxin concentrations in the diluted samples were measured. Results: Endotoxin was detected in both the unused culture medium and the culture supernatant of the epithelial cell sheets at higher concentrations than the criterion value. Therefore, endotoxin concentrations in the reagents used to prepare the culture medium were measured and were found to be below the criterion value, except for cholera toxin. On the other hand, three lots of cholera toxin products were used for the measurement, and the endotoxin concentrations were higher than the criterion value. The results indicate that the endotoxin contamination is caused by the cholera toxin product. Conclusions: To prevent endotoxin contamination in cell-based medicinal products, endotoxin concentrations in reagents used for the fabrication should be measured in the facility conducting clinical research or confirmed by an adequate certificate of analysis from the manufacturers of the reagents.

Early use of oral cholera vaccines as a prime control measure during outbreaks: Necessary but not sufficient.

INTRODUCTION: Despite being a preventable and treatable disease, cholera remains a public health problem in Sudan. The objective of the outbreak investigation was to identify associated risk factors that would help institute appropriate control measures. MATERIAL AND METHODS: A case control study design was chosen to identify the risk factors for cholera in Gadarif State. RESULTS: Multi-variate analysis of identified two risk factors and three preventive factors for cholera in Gadarif City. RISK FACTORS: Buying foods or drinks from street vendors (OR = 71.36), 95 % CI: 16.58-307.14), living in an urban setting (Gadarif City) (OR = 5.38), 95 % CI: 2.10-13.81); and the preventive factors were: Washing hands with water after defecation but without soap (OR = 0.16), 95 % CI: 0.04-0.63) or with soap (OR = 0.01), 95 % CI: 0.00-0.03), washing hands before eating (OR = 0.15), 95 % CI: 0.05-0.51) and taking Oral Cholera Vaccine (OCV) (OR = 0.19, 95 % CI: 0.08-0.44). The effectiveness of OCV (VE) was (Unadjusted VE: 80 %, 95 % CI: 69 %-87 %) or (Adjusted VE = 81.0 %, 95 % CI: 56.0 %-92.0 %). DISCUSSION: Cholera outbreaks, especially in the setting of a complex humanitarian crises, can spread rapidly, resulting in many deaths, and quickly become a public health crisis. Implementation of a community-wide vaccination campaign using OCV as early as possible during the outbreak while implementing other control measures to target hotspots and at-risk populations would expedite halting outbreaks of cholera and save lives.

Assessment of Knowledge of Communicable Diseases Among Medical Students at Al-Balqa Applied University.

INTRODUCTION: Medical education is the foundation of knowledge among medical students. This study aims to investigate the knowledge of medical students at Al-Balqa Applied University, exploring their awareness of five communicable diseases, namely, leishmaniasis, hepatitis B, tuberculosis, measles, and cholera. METHODS: This cross-sectional survey included 271 participants who answered a structured validated questionnaire with varying questions on causes, symptoms, complications, transmission routes, and preventive measures for each disease. RESULTS: Knowledge of all five communicable diseases was low. Leishmaniasis knowledge was notably low (mean=6.07, SD=1.43), with participants grappling with misconceptions about transmission modes, symptoms, and preventability. Hepatitis B knowledge was also low (mean=10.46, SD=1.67), especially regarding transmission modes, revealing that 76% of students were unaware of how the virus spreads. Tuberculosis knowledge unveiled gaps (mean=7.007, SD=1.90), particularly in recognizing the causes, symptoms, and transmission routes. Measles knowledge (mean=9.56, SD=1.92) indicated a robust understanding of symptoms but unveiled misconceptions about complications and transmission routes. For cholera (mean=14.50, SD=1.98), a knowledge of symptoms was demonstrated, but confusion about causative agents, transmission routes, and preventive measures was highlighted. CONCLUSION: The findings of the study emphasize the critical need for enhanced educational strategies including curriculum revisions, increased practical exposure, engaging awareness campaigns, and the integration of interactive learning methods to increase knowledge about communicable diseases.

In Commemoration of Dr. Mostafa Pourtaghva Shahrestani, a Pioneer in Infectious Disease Research.

It is important to honor the contributions of scientific leaders who have dedicated their lives to advancing knowledge and serving their country. One way is to document their experiences and personalities in a documentary format, which can serve as a historical record and an inspiration for future generations. Dr. Mostafa Pourtaghva Shahrestani, a renowned physician and specialist in infectious diseases and tropical medicine, has made significant contributions to public health in Iran. He has played a crucial role in controlling infectious diseases such as smallpox, tuberculosis, rabies, plague, and cholera. Throughout his career, he has held various executive positions, including the head of Pasteur Hospital and the director of the Pasteur Institute of Iran. Dr. Pourtaghva's life is a testament to his unwavering dedication to public health services, as evidenced by his continuous effort, love, and interest in honest work. His inspiring story can serve as a model for those who seek to follow in his footsteps.

Comparison of analysis methods to classify cholera hotspots in Ethiopia from 2015 to 2021.

Cholera continues to represent a major public health concern in Ethiopia. The country has developed a Multi-sectoral National Cholera Elimination Plan in 2022, which targets prevention and control interventions in cholera hotspots. Multiple methods to classify cholera hotspots have been used in several countries. Since 2014, a classification method developed by United Nations Children's Fund has been applied to guide water, sanitation and hygiene interventions throughout Sub-Saharan Africa based on three outbreak parameters: frequency, duration and standardized attack rate. In 2019, the Global Task Force on Cholera Control (GTFCC) proposed a method based on two parameters: average annual cholera incidence and persistence. In 2023, an updated GTFCC method for multisectoral interventions considers three epidemiological indicators (cumulative incidence, cumulative mortality and persistence,) and a cholera-case confirmation indicator. The current study aimed to classify cholera hotspots in Ethiopia at the woreda level (equivalent to district level) applying the three methods and comparing the results to optimize the hotspot targeting strategy. From 2015 to 2021, cholera hotspots were located along major routes between Addis Ababa and woredas adjacent to the Kenya and Somalia borders, throughout Tigray Region, around Lake Tana, and in Afar Region. The multi-method comparison enables decision makers to prioritize interventions according to a sub-classification of the highest-priority areas.

Analyses of drinking water quality during a protracted cholera epidemic in Malawi - a cross-sectional study of key physicochemical and microbiological parameters.

Anecdotal evidence and available literature indicated that contaminated water played a major role in spreading the prolonged cholera epidemic in Malawi from 2022 to 2023. This study assessed drinking water quality in 17 cholera-affected Malawi districts from February to April 2023. Six hundred and thirty-three records were analysed. The median counts/100 ml for thermotolerant coliform was 98 (interquartile range (IQR): 4-100) and that for Escherichia coli was 0 (IQR: 0-9). The drinking water in all (except one) districts was contaminated by thermotolerant coliform, while six districts had their drinking water sources contaminated by E. coli. The percentage of contaminated drinking water sources was significantly higher in shallow unprotected wells (80.0% for E. coli and 95.0% for thermotolerant coliform) and in households (55.8% for E. coli and 86.0% for thermotolerant coliform). Logistic regression showed that household water has three times more risk of being contaminated by E. coli and two and a half times more risk of being contaminated by thermotolerant coliform compared to other water sources. This study demonstrated widespread contamination of drinking water sources during a cholera epidemic in Malawi, which may be the plausible reason for the protracted nature of the epidemic.

Molecular identification of Proteus mirabilis, Vibrio species leading to CRISPR-Cas9 modification of tcpA and UreC genes causing cholera and UTI.

Heavy metal accumulation increases rapidly in the environment due to anthropogenic activities and industrialization. The leather and surgical industry produces many contaminants containing heavy metals. Cadmium, a prominent contaminant, is linked to severe health risks, notably kidney and liver damage, especially among individuals exposed to contaminated wastewater. This study aims to leverage the natural cadmium resistance mechanisms in bacteria for bioaccumulation purposes. The industrial wastewater samples, characterized by an alarming cadmium concentration of 29.6 ppm, 52 ppm, and 76.4 ppm-far exceeding the recommended limit of 0.003 ppm-were subjected to screening for cadmium-resistant bacteria using cadmium-supplemented media with CdCl2. 16S rRNA characterization identified Vibrio cholerae and Proteus mirabilis as cadmium-resistant bacteria in the collected samples. Subsequently, the cadmium resistance-associated cadA gene was successfully amplified in Vibrio species and Proteus mirabilis, revealing a product size of 623 bp. Further analysis of the identified bacteria included the examination of virulent genes, specifically the tcpA gene (472 bp) associated with cholera and the UreC gene (317 bp) linked to urinary tract infections. To enhance the bioaccumulation of cadmium, the study proposes the potential suppression of virulent gene expression through in-silico gene-editing tools such as CRISPR-Cas9. A total of 27 gRNAs were generated for UreC, with five selected for expression. Similarly, 42 gRNA sequences were generated for tcpA, with eight chosen for expression analysis. The selected gRNAs were integrated into the lentiCRISPR v2 expression vector. This strategic approach aims to facilitate precise gene editing of disease-causing genes (tcpA and UreC) within the bacterial genome. In conclusion, this study underscores the potential utility of Vibrio species and Proteus mirabilis as effective candidates for the removal of cadmium from industrial wastewater, offering insights for future environmental remediation strategies.

The Mutagenic Plasticity of the Cholera Toxin B-Subunit Surface Residues: Stability and Affinity.

Mastering selective molecule trafficking across human cell membranes poses a formidable challenge in healthcare biotechnology while offering the prospect of breakthroughs in drug delivery, gene therapy, and diagnostic imaging. The cholera toxin B-subunit (CTB) has the potential to be a useful cargo transporter for these applications. CTB is a robust protein that is amenable to reengineering for diverse applications; however, protein redesign has mostly focused on modifications of the N- and C-termini of the protein. Exploiting the full power of rational redesign requires a detailed understanding of the contributions of the surface residues to protein stability and binding activity. Here, we employed Rosetta-based computational saturation scans on 58 surface residues of CTB, including the GM1 binding site, to analyze both ligand-bound and ligand-free structures to decipher mutational effects on protein stability and GM1 affinity. Complimentary experimental results from differential scanning fluorimetry and isothermal titration calorimetry provided melting temperatures and GM1 binding affinities for 40 alanine mutants among these positions. The results showed that CTB can accommodate diverse mutations while maintaining its stability and ligand binding affinity. These mutations could potentially allow modification of the oligosaccharide binding specificity to change its cellular targeting, alter the B-subunit intracellular routing, or impact its shelf-life and in vivo half-life through changes to protein stability. We anticipate that the mutational space maps presented here will serve as a cornerstone for future CTB redesigns, paving the way for the development of innovative biotechnological tools.

Spatiotemporal dynamics of cholera hotspots in the Democratic Republic of the Congo from 1973 to 2022.

BACKGROUND: Since the early 1970s, cholera outbreaks have been a major public health burden in the Democratic Republic of Congo (DRC). Cholera cases have been reported in a quasi-continuous manner in certain lakeside areas in the Great Lakes Region. As these cholera-endemic health zones constitute a starting point for outbreaks and diffusion towards other at-risk areas, they play a major role in cholera dynamics in the country. Monitoring the spatiotemporal dynamics of cholera hotspots and adjusting interventions accordingly thus reduces the disease burden in an efficient and cost-effective manner. METHODS: A literature review was conducted to describe the spatiotemporal dynamics of cholera in the DRC at the province level from 1973 to 1999. We then identified and classified cholera hotspots at the provincial and health zone levels from 2003 to 2022 and described the spatiotemporal evolution of hotspots. We also applied and compared three different classification methods to ensure that cholera hotspots are identified and classified according to the DRC context. RESULTS: According to all three methods, high-priority hotspots were concentrated in the eastern Great Lakes Region. Overall, hotspots largely remained unchanged over the course of the study period, although slight improvements were observed in some eastern hotspots, while other non-endemic areas in the west experienced an increase in cholera outbreaks. The Global Task Force on Cholera Control (GTFCC) and the Department of Ecology and Infectious Disease Control (DEIDC) methods largely yielded similar results for the high-risk hotspots. However, the medium-priority hotspots identified by the GTFCC method were further sub-classified by the DEIDC method, thereby providing a more detailed ranking for priority targeting. CONCLUSIONS: Overall, the findings of this comprehensive study shed light on the dynamics of cholera hotspots in the DRC from 1973 to 2022. These results may serve as an evidence-based foundation for public health officials and policymakers to improve the implementation of the Multisectoral Cholera Elimination Plan, guiding targeted interventions and resource allocation to mitigate the impact of cholera in vulnerable communities.

Sporadic regional re-emergent cholera: a 19th century problem in the 21st century.

Cholera, caused by Vibrio cholerae, is a severe diarrheal disease that necessitates prompt diagnosis and effective treatment. This review comprehensively examines various diagnostic methods, from traditional microscopy and culture to advanced nucleic acid testing like polymerase spiral reaction and rapid diagnostic tests, highlighting their advantages and limitations. Additionally, we explore evolving treatment strategies, with a focus on the challenges posed by antibiotic resistance due to the activation of the SOS response pathway in V. cholerae. We discuss promising alternative treatments, including low-pressure plasma sterilization, bacteriophages, and selenium nanoparticles. The paper emphasizes the importance of multidisciplinary approaches combining novel diagnostics and treatments in managing and preventing cholera, a persistent global health challenge. The current re-emergent 7th pandemic of cholera commenced in 1961 and shows no signs of abeyance. This is probably due to the changing genetic profile of V. cholerae concerning bacterial pathogenic toxins. Given this factor, we argue that the disease is effectively re-emergent, particularly in Eastern Mediterranean countries such as Lebanon, Syria, etc. This review considers the history of the current pandemic, the genetics of the causal agent, and current treatment regimes. In conclusion, cholera remains a significant global health challenge that requires prompt diagnosis and effective treatment. Understanding the history, genetics, and current treatments is crucial in effectively addressing this persistent and re-emergent disease.

Vibrio cholerae virulence and its suppression through the quorum-sensing system.

Vibrio cholerae is a cholera-causing pathogen known to instigate severe contagious diarrhea that affects millions globally. Survival of vibrios depend on a combination of multicellular responses and adapt to changes that prevail in the environment. This process is achieved through a strong communication at the cellular level, the process has been recognized as quorum sensing (QS). The severity of infection is highly dependent on the QS of vibrios in the gut milieu. The quorum may exist in a low/high cell density (LCD/HCD) state to exert a positive or negative response to control the regulatory pathogenic networks. The impact of this regulation reflects on the transition of pathogenic V. cholerae from the environment to infect humans and cause outbreaks or epidemics of cholera. In this context, the review portrays various regulatory processes and associated virulent pathways, which maneuver and control LCD and HCD states for their survival in the host. Although several treatment options are existing, promotion of therapeutics by exploiting the virulence network may potentiate ineffective antibiotics to manage cholera. In addition, this approach is also useful in resource-limited settings, where the accessibility to antibiotics or conventional therapeutic options is limited.

Congruity of genomic and epidemiological data in modelling of local cholera outbreaks.

Cholera continues to be a global health threat. Understanding how cholera spreads between locations is fundamental to the rational, evidence-based design of intervention and control efforts. Traditionally, cholera transmission models have used cholera case-count data. More recently, whole-genome sequence data have qualitatively described cholera transmission. Integrating these data streams may provide much more accurate models of cholera spread; however, no systematic analyses have been performed so far to compare traditional case-count models to the phylodynamic models from genomic data for cholera transmission. Here, we use high-fidelity case-count and whole-genome sequencing data from the 1991 to 1998 cholera epidemic in Argentina to directly compare the epidemiological model parameters estimated from these two data sources. We find that phylodynamic methods applied to cholera genomics data provide comparable estimates that are in line with established methods. Our methodology represents a critical step in building a framework for integrating case-count and genomic data sources for cholera epidemiology and other bacterial pathogens.

Spatiotemporal distribution of climate-sensitive disease incidences in ethiopia: a longitudinal retrospective analysis of Malaria, Meningitis, Cholera, Dysentery, Leishmaniasis and Dengue fever between 2010 and 2022/2023.

BACKGROUND: Understanding the temporal and geographic distribution of disease incidences is crucial for effective public health planning and intervention strategies. This study presents a comprehensive analysis of the spatiotemporal distribution of disease incidences in Ethiopia, focusing on six major diseases: Malaria, Meningitis, Cholera and Dysentery, over the period from 2010 to 2022, whereas Dengue Fever and Leishmaniasis from 2018 to 2023. METHODS: Using data from Ethiopian public health institute: public health emergency management (PHEM), and Ministry of Health, we examined the occurrence and spread of each disease across different regions of Ethiopia. Spatial mapping and time series analysis were employed to identify hotspots, trends, and seasonal variations in disease incidence. RESULTS: The findings reveal distinct patterns for each disease, with varying cases and temporal dynamics. Monthly wise, Malaria exhibits a cyclical pattern with a peak during the rainy and humid season, while Dysentery, Meningitis and Cholera displays intermittent incidences. Dysentery cases show a consistent presence throughout the years, while Meningitis remains relatively low in frequency but poses a potential threat due to its severity. Dengue fever predominantly occurs in the eastern parts of Ethiopia. A significant surge in reported incident cases occurred during the years 2010 to 2013, primarily concentrated in the Amhara, Sidama, Oromia, Dire Dawa, and Benishangul-Gumuz regions. CONCLUSIONS: This study helps to a better understanding of disease epidemiology in Ethiopia and can serve as a foundation for evidence-based decision-making in disease prevention and control. By recognizing the patterns and seasonal changes associated with each disease, health authorities can implement proactive measures to mitigate the impact of outbreaks and safeguard public health in the region.

PIRES2-EGFP/CTB-UreI vaccination activated a mixed Th1/Th2/Th17 immune system defense towards Helicobacter pylori infection in the BALB/c mice model.

The occurrence of gastritis, gastric ulcers, distal gastric cancer, and gastric mucosal lymphoma in humans is strongly associated with Helicobacter pylori (H. pylori). Vaccination is an effective preventive measure due to the increasing prevalence of antibiotic resistance. Fusion vaccination is a potentially practical approach. A fusion vaccine was created in this study by combining the cholera toxin B subunit (CTB) with the antigenic H. pylori urease I subunit (CTB-UreI). The CTB-UreI DNA vaccine was chemically cloned into pIRES2-EGFP, and the success of the cloning was validated using PCR and restriction enzyme digestion. An investigation was conducted on the induction of CTB-UreI in Escherichia coli BL21(DE3). The immunogenicity and immune-protective efficacy of the vaccination were assessed in BALB/c mice. The Western blot assay successfully identified the activation of CTB-UreI. In comparison, BALB/c mice receiving pIRES2-EGFP/CTB-UreI vaccination exhibited higher IgG, IgA, IFN-γ, IL-4, and IL-17 levels in their blood samples. In addition, there was a decrease in stomach injuries and bacterial loads. Furthermore, BALB/c mice inoculated with pIRES2-EGFP/CTB-UreI showed a high level of immunity (100%) against the H. pylori challenge. The pIRES2-EGFP/CTB-UreI elicited a combination of Th1/Th2/Th17 immune responses, possibly contributing to an effective defence mechanism. Our data suggests that using this fusion vaccine to prevent H. pylori infection is a promising option.

Facility capacity and provider knowledge for cholera surveillance and diarrhoea case management in cholera hotspots in the Democratic Republic of Congo - a mixed-methods study.

BACKGROUND: Wider healthcare-strengthening interventions are recommended in cholera hotspots and could benefit other types of diarrhoeal diseases which contribute to greater mortality than cholera. OBJECTIVE: Describe facility capacity and provider knowledge for case management of diarrhoea and cholera surveillance in cholera hotspots in the Democratic Republic of Congo (DRC) among health facilities, drug shops, and traditional health practitioners. METHODS: We conducted a sequential exploratory mixed-method study, using focus group discussions, facility audits, and provider knowledge questionnaires during September and October 2022 in North Kivu and Tanganyika provinces, Eastern DRC. Content analysis was used for qualitative data. Quantitative data were summarised by facility level and healthcare provider type. Audit and knowledge scores (range 0-100) were generated. Multivariable linear regression estimated association between scores and explanatory factors. Qualitative and quantitative data were triangulated during interpretation. RESULTS: Overall, 244 facilities and 308 providers were included. The mean audit score for health facilities was 51/100 (SD: 17). Private facilities had an -11.6 (95% CI, -16.7 to -6.6) lower adjusted mean score compared to public. Mean knowledge score was 59/100 (95% CI, 57 to 60) for health facility personnel, 46/100 (95% CI, 43 to 48) for drug shop vendors and 37/100 (95% CI, 34 to 39) for traditional health practitioners. Providers had particularly low knowledge concerning when to check for low blood sugar, use of nasogastric tubes, and dosing schedules. Knowledge about case definitions for cholera was similar between groups (range 41-58%) except for traditional health practitioners for the definition during an outbreak 15/73 (21%). CONCLUSIONS: Increasing awareness of cholera case definitions in this context could help improve cholera surveillance and control. Increased support and supervision, especially for private providers, could help ensure facilities are equipped to provide safe care. More nuanced aspects of case management should be emphasised in provider training.

The reactive cholera vaccination campaign in urban Dhaka in 2022: experience, lessons learned and future directions.

Introduction: An upsurge of diarrheal cases occurred in Dhaka, Bangladesh, with approximately 30% of the cases being identified as cholera in 2022. To combat this situation, a reactive Oral Cholera Vaccination campaign was organized in five highly cholera-affected areas of Dhaka city. The paper is a descriptive tale of experience gathering, organization and implementation of reactive oral cholera vaccination campaign. Study design: This is a descriptive report of a reactive oral cholera vaccination campaign. Methods: Population density maps were generated using GIS technology before launching the campaign. The target population comprised individuals aged over one year, excluding pregnant women, totaling 2,374,976 people residing in above mentioned areas. The campaign utilized Euvichol-Plus, an OCV with adherence to the necessary cold chain requirements. Total 700 teams, each consisting of six members, were deployed across the five zones. The campaign was conducted in two rounds, where first round took place in June-July 2022, followed by second round in August 2022. During the campaign, data on adverse events following immunization (AEFI) was collected. Expert teams from various government and non-government organizations monitored regularly and ensured the campaign's success. Results: The first round achieved a coverage rate of 99%, whereas in the second round, 86.3% of individuals among the first dose recipients. During the campaigns, a total of 57 AEFIs were reported. Conclusions: This campaign serves as a model for a multispectral approach in combating cholera epidemics, highlighting the collaborative efforts of policymakers, health authorities, local communities, and health partners.

Experience of caregivers on the continuum of care and prevention of malnutrition among children with cholera in Ethiopia: a phenomenology study.

INTRODUCTION: Malnutrition is a public health problem in low- and middle-income countries among children. Although illnesses such as diarrhea are common immediate drivers of childhood malnutrition, their consequences could be averted through optimal sick child feeding and care to ensure the continuum of care. This study aimed to explore the lived experiences of mothers/caregivers on continuum of care to prevent malnutrition among children with cholera in Ethiopia. METHODS: A phenomenology study design was applied to explore experiences of mothers/caregivers in the Bale and Guji zones of the Oromia region, southeast Ethiopia, from November to December 2022 using an unstructured interview guide. The saturation of ideas was used to stop the in-depth interview. Translated data were cleaned and imported into ATLAS.ti7 software for analysis. Using an open coding system, the data were coded into a meaningful context. Deeper immersion into data with repeated reading, creating themes, subthemes, and family/category were carried out. In coding and categorization, multiple coders were involved. The finding was presented using well-spoken verbatim/quotes as illustrations and in narratives. RESULTS: In this qualitative study, ten participants were taken to explore their lived experience on the continuum of care for children with acute malnutrition and cholera. The study found that poverty, expensive cost of living, and poor utilization of diversified food were challenges. Moreover, health facilities did not provide any services to mothers whose child was admitted for malnutrition treatment. Children five years and above were excluded from both therapeutic food and screening for malnutrition program. Interruptions of supplies, low attention given to child feeding, inadequate knowledge, and lack of time to prepare diversified food were the main findings. CONCLUSION: Poverty, poor feeding habits, supplies interruption and non-inclusion of malnourished children five and above in screening for malnutrition and in the therapeutic feeding program is missed opportunities that lead to decreased early detection and treatment of malnutrition among children with cholera. Moreover, mothers/caregivers did not receive any service from health facilities when their child was admitted for treatment of malnutrition. This situation forces them to stop treatment before their child recovers from malnutrition, which has a negative impact on the continuum of care and prevention of malnutrition. Therefore, we strongly recommend strengthening emergency nutrition within the country's health system and revising the food and nutrition policy to incorporate emergency nutrition, with a particular focus on children under the age of fifteen. Additionally, it is important that the study's recommendations underscore the significance of a multi-sectoral approach that involves collaboration among the health sector, government agencies, and non-governmental organizations. Moreover, adaptive agricultural products be made easily accessible to the community which is crucial in effective preventing and reducing malnutrition in children in the study and similar settings.

Re-evaluating John Snow's 1856 south London study.

John Snow, the London doctor who studied cholera in the 1840s and 1850s, argued in Snow (1856) that water exerted an "overwhelming influence" on mortality in a region of south London during the 1854 outbreak. In a paper re-assessing Snow's analysis, Koch and Denike (2006) claim that "Snow made not merely minor arithmetic errors but more importantly critical, conceptual mistakes that adversely affected his results." The claim of errors and mistakes is incorrect and due to a misreading or misunderstanding of Snow's data and analysis. Koch and Denike apply an inappropriate statistical test to Snow's original data (and do so incorrectly). More importantly, due to the misreading of the historical record they alter the underlying primary-source data, rendering their results invalid. Analysis of the data following Snow's approach but with modern statistical tools strongly supports Snow's claim for the primacy of water in accounting for variation in cholera mortality.