Yeast whole genome sequencing (WGS) lacks end-to-end workflows that identify genetic engineering. Here we present Prymetime, a tool that assembles yeast plasmids and chromosomes and annotates genetic engineering sequences. It is a hybrid workflow—it uses short and long reads as inputs to perform separate linear and circular assembly steps. This structure is necessary to accurately resolve genetic engineering sequences in plasmids and the genome. We show this by assembling diverse engineered yeasts, in some cases revealing unintended deletions and integrations. Furthermore, the resulting whole genomes are high quality, although the underlying assembly software does not consistently resolve highly repetitive genome features. Finally, we assemble plasmids and genome integrations from metagenomic sequencing, even with 1 engineered cell in 1000. This work is a blueprint for building WGS workflows and establishes WGS-based identification of yeast genetic engineering.

Objectives:To undertake an descriptive analysis of the health needs, healthcare practices and barriers to accessing healthcare faced by women in Lower Napo River Region, Peru, and to understand health literacy regarding cervical cancer and the need for more effective cervical cancer screening services.Methods:We performed a community-based needs assessment adapting Demographic and Health survey methodology with additional questions determining female health literacy on cervical cancer and assessing the availability and need for cervical cancer screening services. We surveyed women (N = 121) across all households in six communities along the Lower Napo River, Loreto, Peru, in May 2015. Data were collected as part of the larger Amazon Community Based Participation Cervical Cancer Screen-and-Treat Programme. Survey data were compared to national results from ENDES 2014.Results:Comparison between our findings and the ENDES 2014 survey highlighted considerable inequality between indigenous or mixed indigenous, rural populations in Loreto, Peru, and national population data averages over level of formal education, literacy, barriers to accessing healthcare and maternal and sexual health. Even though only 5.9% (N = 7/117) of women had no formal health insurance coverage, money was reported as the leading barrier accessing healthcare (N = 88/117, 75.2%). Health literacy regarding cervical and breast cancer was poor. A high proportion of women highlighted fear of screening processes (70.8%, N = 80/113) and lack of available services (53.6%, N = 60/112) as barriers to cervical cancer screening.Conclusion:Although progress has been made in improving healthcare access in Peru, such gains have not been experienced equitably and women living in remote communities face persistent marginalization regarding their health. There is a significant need for education related to and screening for cervical cancer in this region that is tailored to the reality of women’s lives in remote communities in Loreto.

No abstract

Introduction In 2016, the WHO published recommendations increasing the number of recommended antenatal care (ANC) visits per pregnancy from four to eight. Prior to the implementation of this policy, coverage of four ANC visits has been suboptimal in many low-income settings. In this study we explore socio-demographic factors associated with early initiation of first ANC contact and attending at least four ANC visits (“ANC4+”) in Malawi using the Malawi Demographic and Health Survey (MDHS) data collected between 2004 and 2016, prior to the implementation of new recommendations. Methods We combined data from the 2004–5, 2010 and 2015–16 MDHS using Stata version 16. Participants included all women surveyed between the ages of 15–49 who had given birth in the five years preceding the survey. We conducted weighted univariate, bivariate and multivariable logistic regression analysis of the effects of each of the predictor variables on the binary endpoint of the woman attending at least four ANC visits and having the first ANC attendance within or before the four months of pregnancy (ANC4+). To determine whether a factor was included in the model, the likelihood ratio test was used with a statistical significance of P< 0.05 as the threshold. Results We evaluated data collected in surveys in 2004/5, 2010 and 2015/6 from 26386 women who had given birth in the five years before being surveyed. The median gestational age, in months, at the time of presenting for the first ANC visit was 5 (inter quartile range: 4–6). The proportion of women initiating ANC4+ increased from 21.3% in 2004–5 to 38.8% in 2015–16. From multivariate analysis, there was increasing trend in ANC4+ from women aged 20–24 years (adjusted odds ratio (aOR) = 1.27, 95%CI:1.05–1.53, P = 0.01) to women aged 45–49 years (aOR = 1.91, 95%CI:1.18–3.09, P = 0.008) compared to those aged 15–19 years. Women from richest socio-economic position ((aOR = 1.32, 95%CI:1.12–1.58, P<0.001) were more likely to demonstrate ANC4+ than those from low socio-economic position. Additionally, women who had completed secondary (aOR = 1.24, 95%CI:1.02–1.51, P = 0.03) and tertiary (aOR = 2.64, 95%CI:1.65–4.22, P<0.001) education were more likely to report having ANC4+ than those with no formal education. Conversely increasing parity was associated with a reduction in likelihood of ANC4+ with women who had previously delivered 2–3 (aOR = 0.74, 95%CI:0.63–0.86, P<0.001), 4–5 (aOR = 0.65, 95%CI:0.53–0.80, P<0.001) or greater than 6 (aOR = 0.61, 95%CI: 0.47–0.79, <0.001) children being less likely to demonstrate ANC4+. Conclusion The proportion of women reporting ANC4+ and of key ANC interventions in Malawi have increased significantly since 2004. However, we found that most women did not access the recommended number of ANC visits in Malawi, prior to the 2016 WHO policy change which may mean that women are less likely to undertake the 2016 WHO recommendation of 8 contacts per pregnancy. Additionally, our results highlighted significant variation in coverage according to key socio-demographic variables which should be considered when devising national strategies to ensure that all women access the appropriate frequency of ANC visits during their pregnancy.

Despite high vaccination rates, the incidence of whooping cough has steadily been increasing in developing countries for several decades. The current acellular pertussis (aP) vaccines all include the major protective antigen pertussis toxin (PTx) and are safer, but they appear to be less protective than infection or older, whole-cell vaccines. To better understand the attributes of individual antibodies stimulated by aP, we isolated plasmablast clones recognizing PTx after booster immunization of two donors. Five unique antibody sequences recognizing native PTx were recovered and expressed as recombinant human IgG1 antibodies. The antibodies all bind different epitopes on the PTx S1 subunit, B oligomer, or S1-B subunit interface, and just one clone neutralized PTx in an in vitro assay. To better understand the epitopes bound by the nonneutralizing S1-subunit antibodies, comprehensive mutagenesis with yeast display provided a detailed map of the epitope recognized by antibodies A8 and E12. Residue R76 is required for antibody A8 binding and is present on the S1 surface but is only partially exposed in the holotoxin, providing a structural explanation for A8's inability to neutralize holotoxin. The B-subunit-specific antibody D8 inhibited PTx binding to a model receptor and neutralized PTx in vitro as well as in an in vivo leukocytosis assay. This is the first study, to our knowledge, to identify individual human antibodies stimulated by the acellular pertussis vaccine and demonstrates the feasibility of using these approaches to address outstanding issues in pertussis vaccinology, including mechanisms of accelerated waning of protective immunity despite repeated aP immunization.