Risk factors for acquisition of meningococcal carriage in the African meningitis belt.

OBJECTIVE: To investigate potential risk factors for acquisition in seven countries of the meningitis belt. METHODS: Households were followed up every 2 weeks for 2 months, then monthly for a further 4 months. Pharyngeal swabs were collected from all available household members at each visit and questionnaires completed. Risks of acquisition over the whole study period and for each visit were analysed by a series of logistic regressions. RESULTS: Over the course of the study, acquisition was higher in: (i) 5-to 14-year olds, as compared with those 30 years or older (OR 3.6, 95% CI 1.4-9.9); (ii) smokers (OR 3.6, 95% CI 0.98-13); and (iii) those exposed to wood smoke at home (OR 2.6 95% CI 1.3-5.6). The risk of acquisition from one visit to the next was higher in those reporting a sore throat during the dry season (OR 3.7, 95% CI 2.0-6.7) and lower in those reporting antibiotic use (OR 0.17, 95% CI 0.03-0.56). CONCLUSIONS: Acquisition of meningococcal carriage peaked in school age children. Recent symptoms of sore throat during the dry season, but not during the rainy season, were associated with a higher risk of acquisition. Upper respiratory tract infections may be an important driver of epidemics in the meningitis belt.

OBJECTIF: Investiguer les facteurs de risque potentiels d'acquisition dans sept pays de la ceinture de la méningite. MÉTHODES: Des ménages ont été suivis toutes les deux semaines pendant deux mois, puis tous les mois pendant quatre mois. Des prélèvements pharyngés sur écouvillons ont été collectés auprès de tous les membres disponibles du ménage à chaque visite et des questionnaires ont été remplis. Les risques d'acquisition sur l'ensemble de la période d'étude et pour chaque visite ont été analysés par une série de régressions logistiques. RÉSULTATS: Au cours de l'étude, l'acquisition a été plus élevée chez: (i) les 5-14 ans, par rapport à ceux âgés de 30 ans ou plus (OR = 3,6; IC95%: 1,4-9,9); (ii) les fumeurs (OR = 3,6; IC95%: 0,98-13); et (iii) les personnes exposées à la fumée de bois à la maison (OR = 2,6; IC95%: 1,3-5,6). Le risque d'acquisition d'une visite à l'autre était plus élevé chez les personnes signalant un mal de gorge pendant la saison sèche (OR = 3,7; IC95%: 2,0-6,7) et plus faible chez celles signalant une utilisation d'antibiotique (OR = 0,17; IC95%: 0,03-0,56). CONCLUSIONS: L'acquisition du portage du méningocoque a culminé chez les enfants d'âge scolaire. Les symptômes récents de maux de gorge pendant la saison sèche, mais pas pendant la saison des pluies, étaient associés à un risque d'acquisition plus élevé. Les infections des voies respiratoires supérieures pourraient être un facteur important d'épidémies dans la ceinture de la méningite.

Hierarchical genomic analysis of carried and invasive serogroup A Neisseria meningitidis during the 2011 epidemic in Chad.

BACKGROUND: Serogroup A Neisseria meningitidis (NmA) was the cause of the 2011 meningitis epidemics in Chad. This bacterium, often carried asymptomatically, is considered to be an "accidental pathogen"; however, the transition from carriage to disease phenotype remains poorly understood. This study examined the role genetic diversity might play in this transition by comparing genomes from geographically and temporally matched invasive and carried NmA isolates. RESULTS: All 23 NmA isolates belonged to the ST-5 clonal complex (cc5). Ribosomal MLST comparison with other publically available NmA:cc5 showed that isolates were closely related, although those from Chad formed two distinct branches and did not cluster with other NmA, based on their MLST profile, geographical and temporal location. Whole genome MLST (wgMLST) comparison identified 242 variable genes among all Chadian isolates and clustered them into three distinct phylogenetic groups (Clusters 1, 2, and 3): no systematic clustering by disease or carriage source was observed. There was a significant difference (p = 0.0070) between the mean age of the individuals from which isolates from Cluster 1 and Cluster 2 were obtained, irrespective of whether the person was a case or a carrier. CONCLUSIONS: Whole genome sequencing provided high-resolution characterization of the genetic diversity of these closely related NmA isolates. The invasive meningococcal isolates obtained during the epidemic were not homogeneous; rather, a variety of closely related but distinct clones were circulating in the human population with some clones preferentially colonizing specific age groups, reflecting a potential age-related niche adaptation. Systematic genetic differences were not identified between carriage and disease isolates consistent with invasive meningococcal disease being a multi-factorial event resulting from changes in host-pathogen interactions along with the bacterium.

Alternative Molecular Methods for Improved Detection of Meningococcal Carriage and Measurement of Bacterial Density.

Conventional methods for detecting pharyngeal carriage of Neisseria meningitidis are complex. There is a need for simpler methods with improved performance. We have investigated two alternative approaches. Three pharyngeal swabs were collected from 999 pupils aged 10 to 18 years in The Gambia. Carriage of N. meningitidis was investigated by using three different methods: (i) plating on Thayer-Martin selective medium and testing by conventional microbiological methods followed by PCR testing; (ii) seeding in Todd-Hewitt broth (THB) and, after culture overnight, testing by PCR; and (iii) compression of the swab on filter paper and, after DNA concentration, testing by PCR. PCR after culture in THB was more than twice as sensitive as conventional methods in detecting N. meningitidis (13.2% versus 5.7%; P < 0.0001). PCR after DNA extraction from filter paper had a sensitivity similar to that of conventional methods (4.9% versus 5.7%; P = 0.33). Capsular genogroups detected by broth culture were genogroups W (21 isolates), B (12 isolates), Y (8 isolates), E (3 isolates), and X (2 isolates), and 68 meningococci had the capsule-null intergenic region. The distributions of genogroups and of capsule-null organisms were similar with each of the three methods. The carriage density in samples extracted from filter paper ranged from 1 to 25,000 DNA copies. PCR of broth cultures grown overnight doubled the yield of N. meningitidis carriage isolates compared with conventional methods. This approach could improve the efficiency of carriage studies. Collection on filter paper followed by quantitative PCR could be useful for density measurement and for carriage studies in areas with limited resources.

Higher Tetanus Toxoid Immunity 2 Years After PsA-TT Introduction in Mali.

BACKGROUND: In 2010, mass vaccination with a then-new meningococcal A polysaccharide-tetanus toxoid protein conjugate vaccine (PsA-TT, or MenAfriVac) was undertaken in 1- to 29-year-olds in Bamako, Mali. Whether vaccination with PsA-TT effectively boosts tetanus immunity in a population with heterogeneous baseline tetanus immunity is not known. We assessed the impact of PsA-TT on tetanus toxoid (TT) immunity by quantifying age- and sex-specific immunity prior to and 2 years after introduction. METHODS: Using a household-based, age-stratified design, we randomly selected participants for a prevaccination serological survey in 2010 and a postvaccination survey in 2012. TT immunoglobulin G (IgG) antibodies were quantified and geometric mean concentrations (GMCs) pre- and postvaccination among all age groups targeted for vaccination were compared. The probability of TT IgG levels ≥0.1 IU/mL (indicating short-term protection) and ≥1.0 IU/mL (indicating long-term protection) by age and sex was determined using logistic regression models. RESULTS: Analysis of 793 prevaccination and 800 postvaccination sera indicated that while GMCs were low pre-PsA-TT, significantly higher GMCs in all age-sex strata were observed 2 years after PsA-TT introduction. The percentage with short-term immunity increased from 57.1% to 88.4% (31.3-point increase; 95% confidence interval [CI], 26.6-36.0;, P < .0001) and with long-term immunity increased from 20.0% to 58.5% (38.5-point increase; 95% CI, 33.7-43.3; P < .0001) pre- and postvaccination. CONCLUSIONS: Significantly higher TT immunity was observed among vaccine-targeted age groups up to 2 years after Mali's PsA-TT mass vaccination campaign. Our results, combined with evidence from clinical trials, strongly suggest that conjugate vaccines containing TT such as PsA-TT should be considered bivalent vaccines because of their ability to boost tetanus immunity.

Continuing effectiveness of serogroup A meningococcal conjugate vaccine, Chad, 2013.

In 2011, vaccination with a serogroup A meningococcal polysaccharide conjugate vaccine was implemented in 3 of 23 regions in Chad. Cases of meningitis declined dramatically in vaccinated areas, but an epidemic continued in the rest of Chad. In 2012, the remaining Chad population was vaccinated, and the epidemic was halted.

Surveillance of Antimicrobial Resistance in the ECOWAS Region: Setting the Scene for Critical Interventions Needed.

Antimicrobial resistance poses a significant challenge to public health globally, leading to increased morbidity and mortality. AMR surveillance involves the systematic collection, analysis, and interpretation of data on the occurrence and distribution of AMR in humans, animals, and the environment for action. The West African Health Organization, part of the Economic Community of West African States (ECOWAS), is committed to addressing AMR in the region. This paper examines the status of AMR surveillance in ECOWAS countries using available WHO data from the TrACSS survey and GLASS enrollments. The analysis reveals that while progress has been made, significant challenges remain. Twelve of the fifteen ECOWAS countries are enrolled in GLASS, and ten have developed national action plans (NAPs) for AMR. However, there is a need to ensure all countries fully implement their NAPs, continue reporting to GLASS, and use the data for evidence-based actions and decision making. Surveillance systems for AMR and antimicrobial consumption/use vary across countries with some demonstrating limited capacity. All countries, except Cabo Verde, reported having a reference laboratory for AMR testing. Strengthening laboratory capabilities, data management and use, and multisectoral coordination are crucial for effective AMR surveillance and response. Based on the findings and the regional context, it is essential to prioritize capacity building, data utilization, and the adoption of standardized guidelines for AMR surveillance. Collaboration among ECOWAS countries, the WAHO, and international partners is essential to address AMR comprehensively. Ensuring a consistent supply of essential antimicrobial medications and reagents is vital.

Timing and source of subtype-C HIV-1 superinfection in the newly infected partner of Zambian couples with disparate viruses.

BACKGROUND: HIV-1 superinfection occurs at varying frequencies in different at risk populations. Though seroincidence is decreased, in the negative partner of HIV-discordant couples after joint testing and counseling in the Zambia Emory HIV Research Project (ZEHRP) cohort, the annual infection rate remains relatively high at 7-8%. Based on sequencing within the gp41 region of each partner's virus, 24% of new infections between 2004 and 2008 were the result of transmission from a non-spousal partner. Since these seroconvertors and their spouses have disparate epidemiologically-unlinked viruses, there is a risk of superinfection within the marriage. We have, therefore, investigated the incidence and viral origin of superinfection in these couples. RESULTS: Superinfection was detected by heteroduplex mobility assay (HMA), degenerate base counting of the gp41 sequence, or by phylogenetic analysis of the longitudinal sequences. It was confirmed by full-length env single genome amplification and phylogenetic analysis. In 22 couples (44 individuals), followed for up to five years, three of the newly infected (initially HIV uninfected) partners became superinfected. In each case superinfection occurred during the first 12 months following initial infection of the negative partner, and in each case the superinfecting virus was derived from a non-spousal partner. In addition, one probable case of intra-couple HIV-1 superinfection was observed in a chronically infected partner at the time of his seroconverting spouse's initial viremia. Extensive recombination within the env gene was observed following superinfection. CONCLUSIONS: In this subtype-C discordant couple cohort, superinfection, during the first year after HIV-1 infection of the previously negative partner, occurred at a rate similar to primary infection (13.6% [95% CI 5.2-34.8] vs 7.8% [7.1-8.6]). While limited intra-couple superinfection may in part reflect continued condom usage within couples, this and our lack of detecting newly superinfected individuals after one year of primary infection raise the possibility that immunological resistance to intra-subtype superinfection may develop over time in subtype C infected individuals.

Inflammatory genital infections mitigate a severe genetic bottleneck in heterosexual transmission of subtype A and C HIV-1.

The HIV-1 epidemic in sub-Saharan Africa is driven largely by heterosexual transmission of non-subtype B viruses, of which subtypes C and A are predominant. Previous studies of subtype B and subtype C transmission pairs have suggested that a single variant from the chronically infected partner can establish infection in their newly infected partner. However, in subtype A infected individuals from a sex worker cohort and subtype B individuals from STD clinics, infection was frequently established by multiple variants. This study examined over 1750 single-genome amplified viral sequences derived from epidemiologically linked subtype C and subtype A transmission pairs very early after infection. In 90% (18/20) of the pairs, HIV-1 infection is initiated by a single viral variant that is derived from the quasispecies of the transmitting partner. In addition, the virus initiating infection in individuals who were infected by someone other than their spouse was characterized to determine if genital infections mitigated the severe genetic bottleneck observed in a majority of epidemiologically linked heterosexual HIV-1 transmission events. In nearly 50% (3/7) of individuals infected by someone other than their spouse, multiple genetic variants from a single individual established infection. A statistically significant association was observed between infection by multiple genetic variants and an inflammatory genital infection in the newly infected individual. Thus, in the vast majority of HIV-1 transmission events in cohabiting heterosexual couples, a single genetic variant establishes infection. Nevertheless, this severe genetic bottleneck can be mitigated by the presence of inflammatory genital infections in the at risk partner, suggesting that this restriction on genetic diversity is imposed in large part by the mucosal barrier.

Human immunodeficiency virus type 1 elite neutralizers: individuals with broad and potent neutralizing activity identified by using a high-throughput neutralization assay together with an analytical selection algorithm.

The development of a rapid and efficient system to identify human immunodeficiency virus type 1 (HIV-1)-infected individuals with broad and potent HIV-1-specific neutralizing antibody responses is an important step toward the discovery of critical neutralization targets for rational AIDS vaccine design. In this study, samples from HIV-1-infected volunteers from diverse epidemiological regions were screened for neutralization responses using pseudovirus panels composed of clades A, B, C, and D and circulating recombinant forms (CRFs). Initially, 463 serum and plasma samples from Australia, Rwanda, Uganda, the United Kingdom, and Zambia were screened to explore neutralization patterns and selection ranking algorithms. Samples were identified that neutralized representative isolates from at least four clade/CRF groups with titers above prespecified thresholds and ranked based on a weighted average of their log-transformed neutralization titers. Linear regression methods selected a five-pseudovirus subset, representing clades A, B, and C and one CRF01_AE, that could identify top-ranking samples with 50% inhibitory concentration (IC(50)) neutralization titers of >or=100 to multiple isolates within at least four clade groups. This reduced panel was then used to screen 1,234 new samples from the Ivory Coast, Kenya, South Africa, Thailand, and the United States, and 1% were identified as elite neutralizers. Elite activity is defined as the ability to neutralize, on average, more than one pseudovirus at an IC(50) titer of 300 within a clade group and across at least four clade groups. These elite neutralizers provide promising starting material for the isolation of broadly neutralizing monoclonal antibodies to assist in HIV-1 vaccine design.

Deciphering human immunodeficiency virus type 1 transmission and early envelope diversification by single-genome amplification and sequencing.

Accurate identification of the transmitted virus and sequences evolving from it could be instrumental in elucidating the transmission of human immunodeficiency virus type 1 (HIV-1) and in developing vaccines, drugs, or microbicides to prevent infection. Here we describe an experimental approach to analyze HIV-1 env genes as intact genetic units amplified from plasma virion RNA by single-genome amplification (SGA), followed by direct sequencing of uncloned DNA amplicons. We show that this strategy precludes in vitro artifacts caused by Taq-induced nucleotide substitutions and template switching, provides an accurate representation of the env quasispecies in vivo, and has an overall error rate (including nucleotide misincorporation, insertion, and deletion) of less than 8 x 10(-5). Applying this method to the analysis of virus in plasma from 12 Zambian subjects from whom samples were obtained within 3 months of seroconversion, we show that transmitted or early founder viruses can be identified and that molecular pathways and rates of early env diversification can be defined. Specifically, we show that 8 of the 12 subjects were each infected by a single virus, while 4 others acquired more than one virus; that the rate of virus evolution in one subject during an 80-day period spanning seroconversion was 1.7 x 10(-5) substitutions per site per day; and that evidence of strong immunologic selection can be seen in Env and overlapping Rev sequences based on nonrandom accumulation of nonsynonymous mutations. We also compared the results of the SGA approach with those of more-conventional bulk PCR amplification methods performed on the same patient samples and found that the latter is associated with excessive rates of Taq-induced recombination, nucleotide misincorporation, template resampling, and cloning bias. These findings indicate that HIV-1 env genes, other viral genes, and even full-length viral genomes responsible for productive clinical infection can be identified by SGA analysis of plasma virus sampled at intervals typical in large-scale vaccine trials and that pathways of viral diversification and immune escape can be determined accurately.

Development of a PCR algorithm to detect and characterize Neisseria meningitidis carriage isolates in the African meningitis belt.

Improved methods for the detection and characterization of carried Neisseria meningitidis isolates are needed. We evaluated a multiplex PCR algorithm for the detection of a variety of carriage strains in the meningitis belt. To further improve the sensitivity and specificity of the existing PCR assays, primers for gel-based PCR assays (sodC, H, Z) and primers/probe for real-time quantitative PCR (qPCR) assays (porA, cnl, sodC, H, E, Z) were modified or created using Primer Express software. Optimized multiplex PCR assays were tested on 247 well-characterised carriage isolates from six countries of the African meningitis belt. The PCR algorithm developed enabled the detection of N. meningitidis species using gel-based and real-time multiplex PCR targeting porA, sodC, cnl and characterization of capsule genes through sequential multiplex PCR assays for genogroups (A, W, X, then B, C, Y and finally H, E and Z). Targeting both porA and sodC genes together allowed the detection of meningococci with a sensitivity of 96% and 89% and a specificity of 78% and 67%, for qPCR and gel-based PCR respectively. The sensitivity and specificity ranges for capsular genogrouping of N. meningitidis are 67% - 100% and 98%-100% respectively for gel-based PCR and 90%-100% and 99%-100% for qPCR. We developed a PCR algorithm that allows simple, rapid and systematic detection and characterisation of most major and minor N. meningitidis capsular groups, including uncommon capsular groups (H, E, Z).

Meningococcal carriage within households in the African meningitis belt: A longitudinal pilot study.

OBJECTIVES: Carriers of Neisseria meningitidis are a key source of transmission. In the African meningitis belt, where risk of meningococcal disease is highest, a greater understanding of meningococcal carriage dynamics is needed. METHODS: We randomly selected an age-stratified sample of 400 residents from 116 households in Bamako, Mali, and collected pharyngeal swabs in May 2010. A month later, we enrolled all 202 residents of 20 of these households (6 with known carriers) and collected swabs monthly for 6 months prior to MenAfriVac vaccine introduction and returned 10 months later to collect swabs monthly for 3 months. We used standard bacteriological methods to identify N. meningitidis carriers and fit hidden Markov models to assess acquisition and clearance overall and by sex and age. RESULTS: During the cross-sectional study 5.0% of individuals (20/400) were carriers. During the longitudinal study, 73 carriage events were identified from 1422 swabs analyzed, and 16.3% of individuals (33/202) were identified as carriers at least once. The majority of isolates were non-groupable; no serogroup A carriers were identified. CONCLUSIONS: Our results suggest that the duration of carriage with any N. meningitidis averages 2.9 months and that males and children acquire and lose carriage more frequently in an urban setting in Mali. Our study informed the design of a larger study implemented in seven countries of the African meningitis belt.

Investigation of correlates of protection against pharyngeal carriage of Neisseria meningitidis genogroups W and Y in the African meningitis belt.

BACKGROUND: Serum bactericidal antibody titres that correlate with protection against invasive meningococcal disease have been characterised. However, titres that are associated with protection against acquisition of pharyngeal carriage of Neisseria meningitidis are not known. METHODS: Sera were obtained from the members of a household in seven countries of the African meningitis belt in which a pharyngeal carrier of N. meningitidis had been identified during a cross-sectional survey. Serum bactericidal antibody titres at baseline were compared between individuals in the household of the carrier who became a carrier of a meningococcus of the same genogroup during six months of subsequent follow-up and household members who did not become a carrier of a meningococcus of this genogroup during this period. RESULTS: Serum bacterial antibody titres were significantly higher in carriers of a serogroup W or Y meningococcus at the time of recruitment than in those who were not a carrier of N. meningitidis of the same genogroup. Serum bactericidal antibody titres to a strain of N. meningitis of the same genogroup as the index cases were no different in individuals who acquired carriage with a meningococcus of the same genogroup as the index case than in those who did not become a carrier during six months of follow-up. CONCLUSION: Serum bacterial antibody titres to N. meningitidis of genogroup W or Y in the range of those acquired by natural exposure to meningococci of these genogroups, or with cross-reactive bacteria, are not associated with protection against acquisition of carriage with meningococci of either of these genogroups.

A Seroepidemiological Study of Serogroup A Meningococcal Infection in the African Meningitis Belt.

The pattern of epidemic meningococcal disease in the African meningitis belt may be influenced by the background level of population immunity but this has been measured infrequently. A standardised enzyme-linked immunosorbent assay (ELISA) for measuring meningococcal serogroup A IgG antibodies was established at five centres within the meningitis belt. Antibody concentrations were then measured in 3930 individuals stratified by age and residence from six countries. Seroprevalence by age was used in a catalytic model to determine the force of infection. Meningococcal serogroup A IgG antibody concentrations were high in each country but showed heterogeneity across the meningitis belt. The geometric mean concentration (GMC) was highest in Ghana (9.09 µg/mL [95% CI 8.29, 9.97]) and lowest in Ethiopia (1.43 µg/mL [95% CI 1.31, 1.57]) on the margins of the belt. The force of infection was lowest in Ethiopia (λ = 0.028). Variables associated with a concentration above the putative protective level of 2 µg/mL were age, urban residence and a history of recent vaccination with a meningococcal vaccine. Prior to vaccination with the serogroup A meningococcal conjugate vaccine, meningococcal serogroup A IgG antibody concentrations were high across the African meningitis belt and yet the region remained susceptible to epidemics.

Pharyngeal carriage of Neisseria species in the African meningitis belt.

OBJECTIVES: Neisseria meningitidis, together with the non-pathogenic Neisseria species (NPNs), are members of the complex microbiota of the human pharynx. This paper investigates the influence of NPNs on the epidemiology of meningococcal infection. METHODS: Neisseria isolates were collected during 18 surveys conducted in six countries in the African meningitis belt between 2010 and 2012 and characterized at the rplF locus to determine species and at the variable region of the fetA antigen gene. Prevalence and risk factors for carriage were analyzed. RESULTS: A total of 4694 isolates of Neisseria were obtained from 46,034 pharyngeal swabs, a carriage prevalence of 10.2% (95% CI, 9.8-10.5). Five Neisseria species were identified, the most prevalent NPN being Neisseria lactamica. Six hundred and thirty-six combinations of rplF/fetA_VR alleles were identified, each defined as a Neisseria strain type. There was an inverse relationship between carriage of N. meningitidis and of NPNs by age group, gender and season, whereas carriage of both N. meningitidis and NPNs was negatively associated with a recent history of meningococcal vaccination. CONCLUSION: Variations in the prevalence of NPNs by time, place and genetic type may contribute to the particular epidemiology of meningococcal disease in the African meningitis belt.

Correction: A Seroepidemiological Study of Serogroup A Meningococcal Infection in the African Meningitis Belt.

[This corrects the article DOI: 10.1371/journal.pone.0147928.].

Field acceptability and effectiveness of the routine utilization of zidovudine to reduce mother-to-child transmission of HIV-1 in West Africa.

OBJECTIVE: To ascertain the field acceptability and effectiveness of the routine utilization of zidovudine in reducing mother-to-child transmission (MTCT) of HIV in breastfed children after a randomized clinical trial demonstrated its efficacy in Côte d'Ivoire and Burkina Faso. METHODS: Pregnant women aged 18 years or older, who had confirmed HIV-1 infection, haemoglobinemia greater than 7 g/dl were enrolled in an open label cohort at 36-38 weeks' gestation to receive an oral short course of zidovudine. Paediatric HIV infection was defined as a positive HIV-1 polymerase chain reaction, or if aged 15 months or older, a positive HIV serology. RESULTS: The acceptability of HIV pretest counselling was significantly higher in the cohort (90.3%) than in the trial (83.7%) (P < 0.001), but the return rate for HIV test results and for inclusion was low. A similar proportion of women accepted starting zidovudine in the cohort, 30.4% compared with 27.3% in the trial (P = 0.13). The proportions of women who took more than 80% of the expected zidovudine regimen were 81.8% before labour, 86.7% during labour, and 88.1% during the postpartum period, compared with those observed during the trial, 78.1, 81.1, and 85%, respectively. The MTCT probability at age 15 months was 19.6% in the cohort (n = 185) versus 21.2% in the trial (P = 0.52). CONCLUSION: The major drawback with the implementation of a short zidovudine regimen to reduce MTCT is HIV counselling and testing procedures. For women who consent, zidovudine is well accepted and efficacious under routine circumstances.