Whole genome sequence of Vibrio cholerae directly from dried spotted filter paper.

BACKGROUND: Global estimates for cholera annually approximate 4 million cases worldwide with 95,000 deaths. Recent outbreaks, including Haiti and Yemen, are reminders that cholera is still a global health concern. Cholera outbreaks can rapidly induce high death tolls by overwhelming the capacity of health facilities, especially in remote areas or areas of civil unrest. Recent studies demonstrated that stool specimens preserved on filter paper facilitate molecular analysis of Vibrio cholerae in resource limited settings. Specimens preserved in a rapid, low-cost, safe and sustainable manner for sequencing provides previously unavailable data about circulating cholera strains. This may ultimately contribute new information to shape public policy response on cholera control and elimination. METHODOLOGY/PRINCIPAL FINDINGS: Whole genome sequencing (WGS) recovered close to a complete sequence of the V. cholerae O1 genome with satisfactory genome coverage from stool specimens enriched in alkaline peptone water (APW) and V. cholerae culture isolates, both spotted on filter paper. The minimum concentration of V. cholerae DNA sufficient to produce quality genomic information was 0.02 ng/μL. The genomic data confirmed the presence or absence of genes of epidemiological interest, including cholera toxin and pilus loci. WGS identified a variety of diarrheal pathogens from APW-enriched specimen spotted filter paper, highlighting the potential for this technique to explore the gut microbiome, potentially identifying co-infections, which may impact the severity of disease. WGS demonstrated that these specimens fit within the current global cholera phylogenetic tree, identifying the strains as the 7th pandemic El Tor. CONCLUSIONS: WGS results allowed for mapping of short reads from APW-enriched specimen and culture isolate spotted filter papers. This provided valuable molecular epidemiological sequence information on V. cholerae strains from remote, low-resource settings. These results identified the presence of co-infecting pathogens while providing rare insight into the specific V. cholerae strains causing outbreaks in cholera-endemic areas

Eff ectiveness of one dose of oral cholera vaccine in response to an outbreak: a case-cohort study

Background Oral cholera vaccines represent a new eff ective tool to fi ght cholera and are licensed as two-dose regimens with 2–4 weeks between doses. Evidence from previous studies suggests that a single dose of oral cholera vaccine might provide substantial direct protection against cholera. During a cholera outbreak in May, 2015, in Juba, South Sudan, the Ministry of Health, Médecins Sans Frontières, and partners engaged in the fi rst fi eld deployment of a single dose of oral cholera vaccine to enhance the outbreak response. We did a vaccine eff ectiveness study in conjunction with this large public health intervention. Methods We did a case-cohort study, combining information on the vaccination status and disease outcomes from a random cohort recruited from throughout the city of Juba with that from all the cases detected. Eligible cases were those aged 1 year or older on the fi rst day of the vaccination campaign who sought care for diarrhoea at all three cholera treatment centres and seven rehydration posts throughout Juba. Confi rmed cases were suspected cases who tested positive to PCR for Vibrio cholerae O1. We estimated the short-term protection (direct and indirect) conferred by one dose of cholera vaccine (Shanchol, Shantha Biotechnics, Hyderabad, India). Findings Between Aug 9, 2015, and Sept 29, 2015, we enrolled 87 individuals with suspected cholera, and an 898-person cohort from throughout Juba. Of the 87 individuals with suspected cholera, 34 were classifi ed as cholera positive, 52 as cholera negative, and one had indeterminate results. Of the 858 cohort members who completed a follow-up visit, none developed clinical cholera during follow-up. The unadjusted single-dose vaccine eff ectiveness was 80·2% (95% CI 61·5–100·0) and after adjusting for potential confounders was 87·3% (70·2–100·0). Interpretation One dose of Shanchol was eff ective in preventing medically attended cholera in this study. These results support the use of a single-dose strategy in outbreaks in similar epidemiological settings

Three transmission events of Vibrio cholerae O1 into Lusaka, Zambia

Cholera has been present and recurring in Zambia since 1977. However, there is a paucity of data on genetic relatedness and diversity of the Vibrio cholerae isolates responsible for these outbreaks. Understanding whether the outbreaks are seeded from existing local isolates or if the outbreaks represent separate transmission events can inform public health decisions. Seventy-two V. cholerae isolates from outbreaks in 2009/2010, 2016, and 2017/2018 in Zambia were characterized using multilocus variable number tandem repeat analysis (MLVA) and whole genome sequencing (WGS). The isolates had eight distinct MLVA genotypes that clustered into three MLVA clonal complexes (CCs). Each CC contained isolates from only one outbreak. The results from WGS revealed both clustered and dispersed single nucleotide variants. The genetic relatedness of isolates based on WGS was consistent with the MLVA, each CC was a distinct genetic lineage and had nearest neighbors from other East African countries. In Lusaka, isolates from the same outbreak were more closely related to themselves and isolates from other countries than to isolates from other outbreaks in other years. Our observations are consistent with i) the presence of random mutation and alternative mechanisms of nucleotide variation, and ii) three separate transmission events of V. cholerae into Lusaka, Zambia. We suggest that locally, case-area targeted invention strategies and regionally, well-coordinated plans be in place to effectively control future cholera outbreaks.https://doi.org/10.1186/s12879-021-06259-

Three doses of COVID-19 mRNA vaccine induce class-switched antibody responses in inflammatory arthritis patients on immunomodulatory therapies

Patients with inflammatory arthritis (IA) are at increased risk of severe COVID-19 due to medication-induced immunosuppression that impairs host defenses. The aim of this study was to assess antibody and B cell responses to COVID-19 mRNA vaccination in IA patients receiving immunomodulatory therapies. Adults with IA were enrolled through the Johns Hopkins Arthritis Center and compared with healthy controls (HC). Paired plasma and peripheral blood mononuclear cell (PBMC) samples were collected prior to and 30 days or 6 months following the first two doses of mRNA vaccines (D2; HC=77 and IA=31 patients), or 30 days following a third dose of mRNA vaccines (D3; HC=11 and IA=96 patients). Neutralizing antibody titers, total binding antibody titers, and B cell responses to vaccine and Omicron variants were analyzed. Anti-Spike (S) IgG and S-specific B cells developed appropriately in most IA patients following D3, with reduced responses to Omicron variants, and negligible effects of medication type or drug withholding. Neutralizing antibody responses were lower compared to healthy controls after both D2 and D3, with a small number of individuals demonstrating persistently undetectable neutralizing antibody levels. Most IA patients respond as well to mRNA COVID-19 vaccines as immunocompetent individuals by the third dose, with no evidence of improved responses following medication withholding. These data suggest that IA-associated immune impairment may not hinder immunity to COVID-19 mRNA vaccines in most individuals

Cholera Hot-Spots and Contextual Factors in Burundi, Planning for Elimination

The Republic of Burundi first reported cholera cases in 1978 and outbreaks have been occurring nearly every year since then. From 2008–2020, 6949 cases and 43 deaths were officially reported. To evaluate Burundi’s potential to eliminate cholera, we identified hotspots using cholera incidence and disease persistence as suggested by the Global Task Force for Cholera Control. The mean annual incidence for each district that reported cholera ranged from 0.29 to 563.14 cases per 100,000 population per year from 2014–2020. Ten of 12 Health Districts which recorded cholera cases reported a mean annual incidence ≥5 per 100,000 for this time period. Cholera cases occur during the second half of the year in the areas near Lake Tanganyika and along the Ruzizi River, with the highest risk district being Bujumbura Centre. Additional research is needed to understand the role of Lake Tanganyika; risks associated with fishing; migration patterns; and other factors that may explain cholera’s seasonality. Due to the consistent epidemiological pattern and the relatively small area affected by cholera, control and elimination are feasible with an integrated program of campaigns using oral cholera vaccine over the short term and community-based interventions including WASH activities for sustained control

Identification of cholera hotspots in Zambia: A spatiotemporal analysis of cholera data from 2008 to 2017.

The global burden of cholera is increasing, with the majority (60%) of the cases occurring in sub-Saharan Africa. In Zambia, widespread cholera outbreaks have occurred since 1977, predominantly in the capital city of Lusaka. During both the 2016 and 2018 outbreaks, the Ministry of Health implemented cholera vaccination in addition to other preventative and control measures, to stop the spread and control the outbreak. Given the limitations in vaccine availability and the logistical support required for vaccination, oral cholera vaccine (OCV) is now recommended for use in the high risk areas ("hotspots") for cholera. Hence, the aim of this study was to identify areas with an increased risk of cholera in Zambia. Retrospective cholera case data from 2008 to 2017 was obtained from the Ministry of Health, Department of Public Health and Disease Surveillance. The Zambian Central Statistical Office provided district-level population data, socioeconomic and water, sanitation and hygiene (WaSH) indicators. To identify districts at high risk, we performed a discrete Poisson-based space-time scan statistic to account for variations in cholera risk across both space and time over a 10-year study period. A zero-inflated negative binomial regression model was employed to identify the district level risk factors for cholera. The risk map was generated by classifying the relative risk of cholera in each district, as obtained from the space-scan test statistic. In total, 34,950 cases of cholera were reported in Zambia between 2008 and 2017. Cholera cases varied spatially by year. During the study period, Lusaka District had the highest burden of cholera, with 29,080 reported cases. The space-time scan statistic identified 16 districts to be at a significantly higher risk of having cholera. The relative risk of having cholera in these districts was significantly higher and ranged from 1.25 to 78.87 times higher when compared to elsewhere in the country. Proximity to waterbodies was the only factor associated with the increased risk for cholera (P<0.05). This study provides a basis for the cholera elimination program in Zambia. Outside Lusaka, the majority of high risk districts identified were near the border with the DRC, Tanzania, Mozambique, and Zimbabwe. This suggests that cholera in Zambia may be linked to movement of people from neighboring areas of cholera endemicity. A collaborative intervention program implemented in concert with neighboring countries could be an effective strategy for elimination of cholera in Zambia, while also reducing rates at a regional level

Evaluation in Cameroon of a Novel, Simplified Methodology to Assist Molecular Microbiological Analysis of V. cholerae in Resource-Limited Settings.

BACKGROUND:Vibrio cholerae is endemic in South Asia and Africa where outbreaks of cholera occur widely and are particularly associated with poverty and poor sanitation. Knowledge of the genetic diversity of toxigenic V. cholerae isolates, particularly in Africa, remains scarce. The constraints in improving this understanding is not only the lack of regular cholera disease surveillance, but also the lack of laboratory capabilities in endemic countries to preserve, store and ship isolates in a timely manner. We evaluated the use of simplified sample preservation methods for molecular characterization using multi-locus variable-number tandem-repeat analysis (MLVA) for differentiation of Vibrio cholerae genotypes. METHODS AND FINDINGS:Forty-seven V. cholerae isolates and 18 enriched clinical specimens (e.g. stool specimens after enrichment in broth) from cholera outbreaks in Cameroon were preserved on Whatman filter paper for DNA extraction. The samples were collected from two geographically distinct outbreaks in the Far North of Cameroon (FNC) in June 2014 and October 2014. In addition, a convenience sample of 14 isolates from the Philippines and 8 from Mozambique were analyzed. All 87 DNAs were successfully analyzed including 16 paired samples, one a cultured isolate and the other the enriched specimen from which the isolate was collected. Genotypic results were identical between 15 enriched specimens and their culture isolates and the other pair differed at single locus. Two closely related, but distinct clonal complexes were identified among the Cameroonian specimens from 2014. CONCLUSIONS:Collecting V. cholerae using simplified laboratory methods in remote and low-resource settings allows for subsequent advanced molecular characterization of V. cholerae O1. These simplified DNA preservation methods identify V. cholerae and make possible timely information regarding the genetic diversity of V. cholerae; our results set the stage for continued molecular epidemiological research to better understand the transmission and dissemination of V. cholerae in Africa and elsewhere worldwide