Predicting Antibiotic Resistance in Gram-Negative Bacilli from Resistance Genes.

We developed a rapid high-throughput PCR test and evaluated highly antibiotic-resistant clinical isolates of Escherichia coli (n = 2,919), Klebsiella pneumoniae (n = 1,974), Proteus mirabilis (n = 1,150), and Pseudomonas aeruginosa (n = 1,484) for several antibiotic resistance genes for comparison with phenotypic resistance across penicillins, cephalosporins, carbapenems, aminoglycosides, trimethoprim-sulfamethoxazole, fluoroquinolones, and macrolides. The isolates originated from hospitals in North America (34%), Europe (23%), Asia (13%), South America (12%), Africa (7%), or Oceania (1%) or were of unknown origin (9%). We developed statistical methods to predict phenotypic resistance from resistance genes for 49 antibiotic-organism combinations, including gentamicin, tobramycin, ciprofloxacin, levofloxacin, trimethoprim-sulfamethoxazole, ertapenem, imipenem, cefazolin, cefepime, cefotaxime, ceftazidime, ceftriaxone, ampicillin, and aztreonam. Average positive predictive values for genotypic prediction of phenotypic resistance were 91% for E. coli, 93% for K. pneumoniae, 87% for P. mirabilis, and 92% for P. aeruginosa across the various antibiotics for this highly resistant cohort of bacterial isolates.

Reported History of Measles and Long-term Impact on Tetanus Antibody Detected in Children 9-59 Months of Age and Receiving 3 Doses of Tetanus Vaccine in the Democratic Republic of the Congo.

BACKGROUND: Recent studies suggest measles-induced immune amnesia could have long-term immunosuppressive effects via preferential depletion of memory CD150+ lymphocytes, and associations with a 2-3 year period of increased mortality and morbidity from infectious diseases other than measles has been shown in children from wealthy and low-income countries. To further examine the associations previous measles virus infection may have on immunologic memory among children in the Democratic Republic of the Congo (DRC), we assessed tetanus antibody levels among fully vaccinated children, with and without a history of measles. METHODS: We assessed 711 children 9-59 months of age whose mothers were selected for interview in the 2013-2014 DRC Demographic and Health Survey. History of measles was obtained by maternal report and classification of children who had measles in the past was completed using maternal recall and measles IgG serostatus obtained from a multiplex chemiluminescent automated immunoassay dried blood spot analysis. Tetanus IgG antibody serostatus was similarly obtained. A logistic regression model was used to identify association of measles and other predictors with subprotective tetanus IgG antibody. RESULTS: Subprotective geometric mean concentration tetanus IgG antibody values were seen among fully vaccinated children 9-59 months of age, who had a history of measles. Controlling for potential confounding variables, children classified as measles cases were less likely to have seroprotective tetanus toxoid antibody (odds ratio: 0.21; 95% confidence interval: 0.08-0.55) compared with children who had not had measles. CONCLUSIONS: History of measles was associated with subprotective tetanus antibody among this sample of children in the DRC who were 9-59 months of age and fully vaccinated against tetanus.

Tetanus seroprotection among children in the Democratic Republic of the Congo, 2013-2014.

BACKGROUND: Tetanus is a potentially fatal disease that is preventable through vaccination. While the Democratic Republic of the Congo (DRC) has continued to improve implementing routine vaccination activities throughout the country, they have struggled to maintain high childhood vaccine coverage. This study aims to examine the seroprevalence of tetanus in children 6 to 59 months to identify areas for intervention and improvement of vaccination coverage. METHODS: In collaboration with the 2013-2014 Demographic and Health Survey, we assessed the seroprevalence of tetanus antibodies among children in the DRC. Dried blood spot samples collected from children 6-59 months of age were processed using a prototype DYNEX Multiplier® chemiluminescent automated immunoassay instrument with a multiplex measles, mumps, rubella, varicella and tetanus assay. Multivariable logistic regression was used to determine factors associated with tetanus vaccination and seroprotection. RESULTS: Overall, 36.1% of children 6-59 months of age reported receiving at least 1 dose of tetanus vaccine while 28.7% reported receiving 3 doses; tetanus seroprotection was 40%. Increasing age in children was associated with decreased tetanus seroprotection, but increased number tetanus vaccinations received. Factors related to increased tetanus seroprotection included number of children in the household, wealth index of the family, urban residence compared to rural, level of maternal education, and province and geography. CONCLUSIONS: Our findings in this nationally representative sample indicate that serology biomarkers may help identify children who are not fully immunized to tetanus more accurately than reported vaccination. While children may be captured for routine immunization activities, as children age, decreasing seroprevalence may indicate additional need to bolster routine vaccination activities and documentation of vaccination in school aged children. Additionally, the study highlights gaps in rural residential areas and vaccination coverage based on maternal education, indicating that policies targeting maternal education and awareness could improve the coverage and seroprevalence of tetanus antibodies in the DRC.

Measles antibody levels among vaccinated and unvaccinated children 6-59 months of age in the Democratic Republic of the Congo, 2013-2014.

BACKGROUND: Measles is endemic in the Democratic Republic of the Congo (DRC), and 89-94% herd immunity is required to halt its transmission. Much of the World Health Organization African Region, including the DRC, has vaccination coverage below the 95% level required to eliminate measles, heightening concern of inadequate measles immunity. METHODS: We assessed 6706 children aged 6-59 months whose mothers were selected for interview in the 2013-2014 DRC Demographic and Health Survey. History of measles was obtained by maternal report, and classification of children who had measles was completed using maternal recall and measles immunoglobulin G serostatus obtained from a multiplex chemiluminescent automated immunoassay dried blood spot analysis. A logistic regression model was used to identify associations of covariates with measles and seroprotection, and vaccine effectiveness (VE) was calculated. RESULTS: Out of our sample, 64% of children were seroprotected. Measles vaccination was associated with protection against measles (OR: 0.15, 95% CI: 0.03, 0.81) when administered to children 12 months of age or older. Vaccination was predictive of seroprotection at all ages. VE was highest (88%) among children 12-24 months of age. CONCLUSION: Our results demonstrated lower than expected seroprotection against measles among vaccinated children. Understanding the factors that affect host immunity to measles will aid in developing more efficient and effective immunization programs in DRC.

Field Test and Validation of the Multiplier Measles, Mumps, Rubella, and Varicella-Zoster Multiplexed Assay System in the Democratic Republic of the Congo by Using Dried Blood Spots.

Here we describe baseline validation studies and field performance of a research-use-only chemiluminescent multiplex serology panel for measles, mumps, rubella, and varicella-zoster virus used with dried blood spots in support of the 2013-2014 Democratic Republic of the Congo Demographic and Health Survey. Characterization of the panel using U.S. FDA-cleared commercial kits shows good concordance for measles, mumps, rubella, and varicella-zoster with average sensitivity across assays of 94.9% and an average specificity of 91.4%. As expected, performance versus available standards validated for plaque-reduction assays does not provide a 1:1 correspondence with international units and yet demonstrates excellent linearity (average Hill's slope = 1.02) and ∼4 logs of dynamic range. In addition, for the four assays, the multiplexed format allowed for inclusion of three positive and two negative controls for each sample. A prototype Dynex Multiplier chemiluminescent automated immunoassay instrument with a charge-coupled device camera provided a rugged and robust processing and data acquisition platform. Performance of a multiplex instrument for serological testing in a substantially resource-limited environment shows excellent reproducibility, minimal cross-reactivity, and a clear discrimination between specific assays and should be considered a viable option for future serosurveys.IMPORTANCE The critical evaluation of immunization programs is key to identifying areas of suboptimal vaccination coverage, monitoring activities, and aiding development of public health policy. For evaluation of vaccine effectiveness, direct antibody binding assay methods, including enzyme immunoassay, enzyme-linked fluorescence assays, and indirect immunofluorescence assay, are most commonly used for detection of IgG antibodies. However, despite their well-demonstrated, reliable performance, they can be labor-intensive and time-consuming and require separate assays for each individual marker. This necessitates increased sample volumes, processing time, and personnel, which may limit assessment to a few key targets in resource-limited settings, that is, low- and middle-income locations where funding for public health or general infrastructure that directly impacts public health is restricted, limiting access to equipment, infrastructure, and trained personnel. One solution is a multiplexed immunoassay, which allows for the detection of multiple analytes in a single reaction for increased efficiency and rapid surveillance of infectious diseases in limited-resource settings. Thus, the scope of the project precluded a full validation, and here we present abbreviated validation studies demonstrating adequate sensitivity, specificity, and reproducibility.

Association of Previous Measles Infection With Markers of Acute Infectious Disease Among 9- to 59-Month-Old Children in the Democratic Republic of the Congo.

BACKGROUND: Transient immunosuppression and increased susceptibility to other infections after measles infection is well known, but recent studies have suggested the occurrence of an "immune amnesia" that could have long-term immunosuppressive effects. METHODS: We examined the association between past measles infection and acute episodes of fever, cough, and diarrhea among 2350 children aged 9 to 59 months whose mothers were selected for interview in the 2013-2014 Democratic Republic of the Congo (DRC) Demographic and Health Survey (DHS). Classification of children who had had measles was completed using maternal recall and measles immunoglobulin G serostatus obtained via dried-blood-spot analysis with a multiplex immunoassay. The association with time since measles infection and fever, cough, and diarrhea outcomes was also examined. RESULTS: The odds of fever in the previous 2 weeks were 1.80 (95% confidence interval [CI], 1.25-2.60) among children for whom measles was reported compared to children with no history of measles. Measles vaccination demonstrated a protective association against selected clinical markers of acute infectious diseases. CONCLUSION: Our results suggest that measles might have a long-term effect on selected clinical markers of acute infectious diseases among children aged 9 to 59 months in the DRC. These findings support the immune-amnesia hypothesis suggested by others and underscore the need for continued evaluation and improvement of the DRC's measles vaccination program.

Prevalence of Rubella Antibodies Among Children in the Democratic Republic of the Congo.

BACKGROUND: While generally mild in children, rubella infection in early pregnancy can lead to miscarriage, fetal death or congenital rubella syndrome. Rubella vaccination is not yet available as a part of routine immunization in the Democratic Republic of the Congo (DRC), and the burden of infection is unknown. METHODS: In collaboration with the 2013-2014 DRC Demographic and Health Survey, a serosurvey was carried out to assess population immunity to vaccine-preventable diseases. Dry blood spot samples collected from children 6-59 months of age were processed using the Dynex Technologies Multiplier FLEX chemiluminescent immunoassay platform (Dynex Technologies, Chantilly, VA). RESULTS: Among the 7195 6- to 59-month-old children, 33% were positive and <1% indeterminate for rubella antibodies in weighted analyses. Seroprevalence was positively associated with age of the child and province, with seropositivity highest in Bandundu (53%) and lowest in Kasai-Oriental (20%). In multivariate analyses, serologic evidence of infection was associated with age of the mother and child, socioeconomic status and geographic location. CONCLUSIONS: Rubella infection is prevalent among children in the DRC, and while most seroconversion occurs in young children, a significant proportion of children remain at risk and may enter reproductive age susceptible to rubella infection. While not currently in place, implementation of a surveillance program will provide improved estimates of both rubella virus circulation and the burden of congenital rubella syndrome. Such information will play an important role in future policy decisions, vaccine delivery strategies and may provide a basis upon which the effectiveness of rubella antigen introduction may be assessed.

Low Varicella Zoster Virus Seroprevalence Among Young Children in the Democratic Republic of the Congo.

BACKGROUND: Varicella zoster virus (VZV) causes both varicella (chickenpox) and herpes zoster (shingles) and is associated with significant global morbidity. Most epidemiological data on VZV come from high-income countries, and to date there are limited data on the burden of VZV in Africa. METHODS: We assessed the seroprevalence of VZV antibodies among children in the Democratic Republic of Congo in collaboration with the 2013-2014 Demographic and Health Survey. Dried blood spot samples collected from children 6-59 months of age were run on Dynex™ Technologies Multiplier FLEX® chemiluminescent immunoassay platform to assess serologic response. Multivariate logistic regression was then used to determine risk factors for VZV seropositivity. RESULTS: Serologic and survey data were matched for 7,195 children 6-59 months of age, among whom 8% were positive and 2% indeterminate for VZV antibodies in weighted analyses. In multivariate analyses, the odds of seropositivity increased with increasing age, increasing socioeconomic status, mother's education level, rural residence, and province (South Kivu, North Kivu, Bandundu, Bas Congo had the highest odds of a positive test result compared with Kinshasa). CONCLUSION: Our data suggest that VZV is circulating in DRC, and seropositivity is low among children 6-59 months. Seropositivity increased with age and varied by other sociodemographic factors, such as geographic location. This study provides the first nationally representative estimates of VZV infection among children in the DRC.

Evidence of Mumps Infection Among Children in the Democratic Republic of Congo.

BACKGROUND: Mumps is an acute viral infection and while the infection is usually mild, complications can lead to permanent sequelae including brain damage and deafness. The burden of mumps is currently unknown the Democratic Republic of Congo (DRC), we therefore assessed susceptibility to mumps infection among children 6-59 months of age. METHODS: In collaboration with the 2013-2014 DRC Demographic and Health Survey, we conducted a serosurvey to assess population immunity to vaccine preventable diseases. Dried blood spot samples were collected from children 6 to 59 months of age and processed at the UCLA-DRC laboratory in Kinshasa, DRC using the Dynex Technologies Multiplier FLEX chemiluminescent immunoassay platform (Dynex multiplex assay, Chantilly, VA). Logistic multivariate analyses were used to determine risk factors for mumps seropositivity. RESULTS: Serologic and survey data were matched for 7195, 6-59 month-old children, among whom 22% were positive and 3% indeterminate for mumps antibodies in weighted analyses. In multivariate analyses, the odds of seropositivity increased with increasing age, female gender, number of children in household, increasing socioeconomic status and province (Kinshasa with the highest odds of positive test result compared with all other provinces). CONCLUSION: These data suggest that mumps virus is circulating in DRC and risk of exposure increases with age. At present, the introduction of a combined measles-mumps-rubella vaccine remains unlikely, as the capacity to maintain adequate vaccine coverage levels for routine immunization must be improved before additional antigens can be considered for the routine immunization schedule.