This page provides an independent statistical analysis and educational interpretation of publicly reported results. ClinicalTrials.gov provides the official trial registry record. Numerical trial results on this page are restricted to the results posted in the ClinicalTrials.gov record; calculations added by Clinical Biostats are labelled as such.
1. Trial at a Glance
This page covers the registry results for Clinical Investigation Study to Evaluate the Consistency and Reproducibility of Two Consecutive Mosquito Feeding Assays (NCT04666350), a registered phase not stated study. The primary outcome is Oocyst Prevalence.
| Feature | Registry value |
|---|---|
| ClinicalTrials.gov identifier | NCT04666350 |
| Phase | Phase not stated |
| Condition | Malaria |
| Allocation | Not Stated |
| Design model | Not Stated |
| Masking | Not Stated |
| Enrollment | 42 |
| Arms in the primary outcome | n/a |
| Lead sponsor | PATH |
| Results posted | Yes |
2. Clinical Question
Population
Adults with varying Plasmodium falciparum gametocyte densities were enrolled as study volunteers. The registry lists an enrollment of 42. The primary outcome population is participants with available data; one participant without surviving mosquitoes in any assay was excluded from that analysis.
Intervention
Participants underwent direct skin feeding assays (DSFA) on Day 1 and Day 2, with the second assay within 24 hours of the first. The registry also lists direct skin feeding assay, primaquine, and artemether/lumefantrine as interventions. The complete treatment details are not reported in the registry record.
Comparator
The study compares assay measurements from the two consecutive feeding days rather than describing a randomized treatment comparison. The registry identifies one Study Volunteers group; a separate control group and allocation procedure are not reported in the registry record.
Primary question
How consistent and reproducible is oocyst prevalence in surviving mosquitoes when DSFA is performed on Day 1 and Day 2? Oocyst prevalence is the percentage of surviving mosquitoes in a cup with at least one oocyst detected in the midgut. Numerical results describing the relationship between the two days are not reported in the registry record.
The question is about repeatability of an assay under closely spaced testing conditions. It is not, on the available record, a comparison of the clinical effectiveness of primaquine or artemether/lumefantrine. The registry’s stated purpose is to assess consistency and reproducibility of two consecutive DSFA procedures in adults with varying gametocyte densities. It also lists oocyst density, sporozoite prevalence, and sporozoite density as secondary outcomes. The record does not report how consistency was defined statistically or which measure was intended to summarize agreement between assay days.
3. Trial Design
This completed clinical investigation enrolled 42 study volunteers. The registry gives the start date as 2021-03-09 and the primary completion date as 2021-12-20. Its stated design objective was to assess the consistency and reproducibility of two consecutive direct skin feeding assays at a 24-hour interval in adults with varying Plasmodium falciparum gametocyte densities.
Participants provided a blood sample on Day 1 for DMFA testing and then took part in a mosquito feeding assay. On Day 2, they underwent a second DMFA and DSFA assay within 24 hours of the first assays. The arm description says participants received one dose of primaquine after completing the Day 2 DSFA, but the available description ends mid-sentence. The full dosing details, the role of artemether/lumefantrine, and any other protocol procedures are not reported in the registry record.
The primary outcome was oocyst prevalence in surviving mosquitoes, assessed 9 days after feeding for the Day 1 and Day 2 assays. The registry identifies a single Study Volunteers group, not separate randomized groups. Allocation is not stated, and the record explicitly does not describe a like-for-like control comparison. The detailed protocol, blinding procedures, assay implementation requirements, and statistical analysis plan are not reported in the registry record.
The registry labels the study completed, but completion status alone does not provide evidence about the consistency of assay results. The numerical outcome results needed to assess that question are not reported in the registry record. The enrollment count is therefore a description of the study, not a substitute for the outcome estimates or their uncertainty.
4. Participant Flow and Accounting
The registry reports how participants moved through each stage of the study. All counts below are exactly as posted; missing counts are not inferred from other stages.
Before assignment: Four hundred participants consented and underwent screening. After informed consent was obtained participants were tested for the presence of mature stage malarial parasites (called gametocytes) in their blood. Forty-two participants met the inclusion criteria and were gametocyte positive and agreed to participate in the mosquito feeding assays.
Overall Study
| Stage | Study Volunteers |
|---|---|
| STARTED | 42 |
| COMPLETED | 42 |
| NOT COMPLETED | 0 |
Analysis population for the primary outcome, as defined in the registry: Participants with available data; one participant did not have surviving mosquitoes in any of the assays and is not included in the analysis. The number of mosquitoes analyzed reflects the number of surviving mosquitoes after 9 days for each feeding assay and time point.
5. Baseline Characteristics
The table reproduces the baseline characteristics posted in the registry, which describe the participants analysed at baseline and help show whether the randomized groups were comparable.
| Characteristic | Unit | Study Volunteers |
|---|---|---|
| Participants analysed | 42 | |
| Age, Continuous | years | 33.4 (SD 9.3) |
| Sex: Female, Male: Female | Participants | 22 |
| Sex: Female, Male: Male | Participants | 20 |
| Race and Ethnicity Not Collected | Participants | — |
| Region of Enrollment: Kenya | participants | 42 |
| Ever had malaria?: Yes | Participants | 42 |
| Ever had malaria?: No | Participants | 0 |
6. Primary Endpoint
The registry lists the primary outcome as Oocyst Prevalence, measured over Feeding assays were performed on Day 1 and Day 2; Oocyst prevalence in surviving mosquitoes was assessed 9 days after feeding (Days 9 and 10)..
| Element | Registry entry |
|---|---|
| Result type | Mean |
| Measure of spread | Standard Deviation |
| Unit | percentage of mosquitoes |
| Endpoint classification | Continuous Endpoint |
The primary endpoint is oocyst prevalence, defined as the percentage of surviving mosquitoes in a cup with at least one oocyst detected in the midgut. The endpoint concerns whether infection was detected in each surviving mosquito, summarized as a percentage for the cup. It is not the number of oocysts in an individual mosquito; oocyst density is listed separately as a secondary outcome.
After feeding, mosquitoes were maintained in locked environmental chambers for 9 days to allow oocyst development. Their midguts were then dissected, stained with 1% mercurochrome, and examined by optical microscopy. An oocyst is described as a structure on the outer wall of the infected mosquito’s stomach that contains developing sporozoites. The endpoint was assessed in surviving mosquitoes, and the number analyzed depended on how many survived for each feeding assay and time point.
Using prevalence makes the endpoint a measure of how often detectable oocysts occurred among surviving mosquitoes in a cup. In general, this kind of binary-status summary can support comparison of assay results, while not conveying the number of oocysts in mosquitoes that were positive. The registry identifies Day 1 and Day 2 assessments, but numerical prevalence estimates and a measure of agreement between them are not reported in the registry record.
7. Why No Re-analysis Is Shown
8. Statistical Interpretation
The registry record establishes the study’s measurement question and describes how the primary outcome was defined. It does not report numerical oocyst-prevalence results for Day 1 or Day 2, nor does it report an estimated difference, association, or agreement measure between the two assays. As a result, the magnitude and precision of assay consistency cannot be judged from the registry record.
Consistency and reproducibility are related but not interchangeable ideas. In general, consistency concerns whether repeated measurements give comparable results, while reproducibility asks whether results can be obtained again under the specified testing conditions. A comparison of assay-day averages alone may not capture how well the measurements correspond for individual participants or cups. The registry record does not state which aspect was emphasized or what statistical criterion would count as adequate consistency.
The available outcome description identifies the denominator as surviving mosquitoes in the same cup that underwent feeding. The number of mosquitoes analyzed reflects survival after 9 days for each assay and time point. Thus the outcome is conditional on survival and on available data, rather than a summary of every mosquito initially exposed. One participant did not have surviving mosquitoes in any assay and was excluded from the primary analysis population. The record does not report the number of surviving mosquitoes for each cup or time point, or the number of participants contributing data to each reported outcome.
No confidence intervals, p-values, or other inferential results for the primary endpoint are reported in the registry record. A confidence interval, in general, describes uncertainty around an estimate and the range of values compatible with the observed data under the stated method. Without an estimate and its uncertainty, readers cannot determine whether apparent differences across assay days are small or large relative to sampling variability. Likewise, a completed status is not evidence that the assays agreed.
The registry lists secondary outcomes of oocyst density, sporozoite prevalence, and sporozoite density, but their numerical findings are not reported in the registry record. No conclusions about those outcomes can be drawn from their names alone. The intervention list includes primaquine and artemether/lumefantrine, but the design description does not establish a randomized treatment comparison. Therefore the record does not support attributing any assay result to either drug.
Overall, the posted information describes the intended repeat-assay evaluation, the endpoint, and an analysis-population exclusion. It does not provide the outcome estimates needed to determine the observed level of consistency, the precision of that assessment, or whether the study met a prespecified reproducibility criterion. Such a criterion and the statistical plan are not reported in the registry record.
9. Statistical Methods Explained
The registry describes a repeated assay assessment, but does not report the statistical analysis plan or the numerical outcome results. The concepts below explain how such data are generally interpreted; they do not identify methods used in this study unless the registry record states them.
What does an assay repeatability analysis ask?
In general, repeatability analysis asks whether repeated measurements made under similar conditions produce comparable values. For this study, the relevant measurements come from DSFA performed on Day 1 and Day 2, within a 24-hour interval. A complete analysis could describe the distribution on each day and quantify how closely paired results correspond. The registry record does not report which summary or agreement statistic was planned or used.
Comparing average values across days is not the same as evaluating agreement for individual paired observations. In general, two sets of measurements can have similar averages while differing substantially from one participant or cup to another. Conversely, a systematic shift between days may be important even when the rank ordering of observations is similar. The reported endpoint definition makes the cup-level percentage the measurement of interest, but the record does not state the unit used for the primary statistical analysis or how clustering was handled.
What does the outcome summarize?
Oocyst prevalence is the percentage of surviving mosquitoes in a cup with at least one detected oocyst. It summarizes presence or absence, not the number of oocysts in a positive mosquito. General statistical interpretation should therefore distinguish prevalence from density outcomes. Oocyst density, sporozoite prevalence, and sporozoite density are listed as secondary outcomes, but their definitions and analysis methods are not reported in the registry record.
The denominator is limited to surviving mosquitoes. In general, when the number observed in a denominator varies, percentages may have different precision across cups. A percentage based on fewer surviving mosquitoes can be less stable than one based on more observations. The registry says that mosquito counts reflect survival after 9 days for each assay and time point, but does not provide those counts or describe any weighting or variance method. The consequences of survival differences for the primary analysis therefore cannot be assessed from the record.
Why does the stated mean and standard deviation matter?
The primary outcome is listed as a continuous measure summarized by a mean and standard deviation. In general, a mean describes the central value of observations, while a standard deviation describes their spread. Because prevalence is expressed as a percentage, the suitability of a mean-based summary depends on the distribution and the analysis unit. The registry does not report the distribution, transformation, model, or alternative summaries considered. It is not possible to infer from the outcome label alone which assumptions were checked or whether they were appropriate.
For repeated measurements, a method that accounts for the paired nature of the Day 1 and Day 2 observations may be relevant. General examples include paired summaries or models that account for within-participant or within-cup dependence. Mosquito-level measurements from one cup are not necessarily independent observations, since mosquitoes in a cup share the same feeding context. The registry record does not state whether the analysis used participant-level, cup-level, or mosquito-level data, or how dependence was addressed.
How should uncertainty be read?
In general, an estimate describes the observed result in the analyzed data, while a confidence interval communicates uncertainty around that estimate under an analysis model. A narrow interval suggests greater precision than a wide interval, but precision does not by itself establish that the repeated assays agree sufficiently for a particular purpose. A decision about acceptable reproducibility would require a criterion that is specified in advance. No such criterion, confidence interval, or primary-outcome estimate is reported in the registry record.
A p-value, in general, assesses how compatible data are with a specified null hypothesis under the assumptions of a statistical test. It does not measure the size of an effect or prove agreement. The registry does not report a p-value or the hypothesis used for the primary outcome. Nor does it report whether the primary analysis was adjusted for gametocyte density, or whether any other covariates were included.
What can planning calculations establish?
General sample-size planning uses assumptions about the target effect, variability, allocation, and statistical method. Planning illustrations are not observed power and do not demonstrate whether an assay was consistent in the completed study. The registry record does not report the trial’s own sample-size justification. It also does not report interim analyses or the analysis plan. Any conclusions about these features would be speculative.
In summary, the registry supports an explanation of the endpoint and repeated-measurement setting, but not a reconstruction of the analysis. The specific statistical methods, treatment of dependence, missing-data procedures, inferential results, and prespecified standard for reproducibility are not reported in the registry record.
10. Multiplicity, Missing Data and Analysis Populations
The registry lists one primary outcome, oocyst prevalence, and three secondary outcomes: oocyst density, sporozoite prevalence, and sporozoite density. It reports four outcome measures and six analyses, but does not explain how those analyses map to outcomes, time points, or comparisons. The analysis plan and any adjustment for multiple testing are not reported in the registry record.
In general, testing several outcomes or comparisons can increase the chance of at least one apparently notable result when each test is considered separately. Methods for addressing multiplicity depend on the study’s inferential goals and prespecified analysis plan. Without that plan and the results, it is not possible to assess whether multiplicity was addressed or how it should affect interpretation.
The primary population is described as participants with available data. One participant did not have surviving mosquitoes in any assay and was excluded from the analysis. The number of mosquitoes analyzed depended on survival after 9 days at each assay and time point. These details identify sources of unavailable outcome data, but do not establish how other missing observations were handled. The registry record does not report whether missingness occurred for other reasons, whether any imputation or sensitivity analysis was used, or whether the analysis population differed across outcomes.
In general, missing-data handling matters when the probability of an observation being unavailable is related to the outcome or to participant characteristics. Here, survival determines which mosquitoes contribute to prevalence, and one participant had no surviving mosquitoes in any assay. The record does not provide enough detail to assess how this selection affects the interpretation of the estimates. It also does not report a formal definition of the analysis population beyond available data or explain how incomplete paired observations, if any, were treated.
The registry does not report a prespecified statistical analysis plan, a multiplicity strategy, or sensitivity analyses. Consequently, neither the robustness of any reported inference nor the impact of missing data can be evaluated from the registry record.
11. Safety Results
The registry reports serious adverse events as affected participants divided by participants at risk.
| Arm | Participants with serious adverse events | Participants at risk | Percentage |
|---|---|---|---|
| Study Volunteers | 0 | 42 | 0.0% |
Deaths
| Arm | Deaths (participants affected of at risk) |
|---|---|
| Study Volunteers | 0 of 42 |
Other (non-serious) adverse events
Participants with at least one non-serious event at or above the registry's 0% reporting threshold.
| Arm | Participants affected of at risk |
|---|---|
| Study Volunteers | 32 of 42 |
Most frequent non-serious adverse event terms
| Event term | Organ system | Study Volunteers |
|---|---|---|
| Pruritus | Skin and subcutaneous tissue disorders | 26 of 42 |
| Urticaria | Skin and subcutaneous tissue disorders | 5 of 42 |
| Abdominal pain upper | Gastrointestinal disorders | 1 of 42 |
| Tonsillitis | Infections and infestations | 1 of 42 |
| Arthropod bite | Injury, poisoning and procedural complications | 1 of 42 |
The registry lists 42 study volunteers at risk for serious adverse events and reports 0 affected by serious adverse events. It also reports 42 at risk for other adverse events and 32 affected by other adverse events. Deaths are reported as 0. No serious adverse event terms are listed.
These figures describe the safety reporting in the registry record. The count of participants affected by other adverse events does not convey the number of events, their severity, timing, duration, or relationship to an intervention. Those details are not reported in the registry record. The absence of listed serious adverse events should be read as the recorded count for this study, not as proof that a treatment or procedure has no risk.
The study is described as a single Study Volunteers group, not as a randomized comparison. There is no separate control group in the record against which to compare adverse-event counts. The intervention list includes DSFA, primaquine, and artemether/lumefantrine, but the safety figures are not broken down by intervention or exposure in the registry record. These counts therefore do not establish comparative safety or identify which procedure or intervention was associated with an event.
More detailed safety interpretation would require event descriptions, timing, exposure information, and an analysis appropriate to the study design. Those details are not reported in the registry record.
12. What the Result Does, and Does Not, Mean
The registry record describes a completed study designed to examine whether oocyst-prevalence measurements from two DSFA procedures performed 24 hours apart are consistent. It specifies the mosquito-based endpoint, the assessment timing, and the fact that the analysis excluded one participant who had no surviving mosquitoes in any assay. These registry entries clarify what the study set out to measure and which participants were included in the stated primary population.
The numerical primary-outcome results are not reported in the registry record. There are no Day 1 or Day 2 prevalence estimates, no measure of agreement, and no confidence interval or p-value. Therefore, the observed consistency and its precision cannot be determined from the registry information. The study’s completed status does not answer the outcome question by itself.
The record also does not specify a threshold for acceptable reproducibility. In general, whether an assay is sufficiently consistent depends on the intended use and on a criterion chosen before interpreting the data. A result that appears similar on average could still show variation across participants or cups, while a quantified measure of agreement would help characterize that variation. Neither the relevant criterion nor such a measure is reported in the registry record.
The listed secondary outcomes broaden the planned measurement scope, but their numerical findings are not reported. The intervention list and the post-Day 2 primaquine description do not turn the study into a randomized treatment comparison. Allocation is not stated, and the record does not describe a like-for-like control group. It is therefore not possible to infer a treatment effect from the study description.
The defensible conclusion is limited: the registry documents an assay-repeatability objective and the way the primary endpoint was defined, but does not provide the results required to establish how well the two assays agreed. It also does not provide enough statistical detail to assess the analysis strategy or missing-data impact.
13. Why This Trial Matters Statistically
Repeated biological assays can vary because of the measurement procedure, the biological material being measured, and ordinary sampling variability. Evaluating whether an assay gives comparable results on closely spaced occasions is therefore a distinct statistical question from testing whether a treatment changes a clinical outcome. This study’s stated focus on two consecutive DSFA procedures makes assay consistency the central issue.
The primary endpoint is based on the proportion of surviving mosquitoes in a cup with at least one oocyst detected. That definition makes the observation unit and denominator important. Survival after feeding affects which mosquitoes contribute, and the number analyzed can vary across assays and time points. Statistical interpretation needs to account for that structure rather than treating each percentage as if it necessarily had equal precision.
The registry’s description also highlights why a reproducibility claim needs more than a label such as completed. A useful assessment generally requires paired results, an agreement measure, uncertainty around that measure, and a prespecified view of what level of variation is acceptable. The record does not report those results or the standard used to judge reproducibility. This makes the distinction between study purpose and demonstrated finding especially important.
The available information is also a reminder that an endpoint definition is not an analysis plan. The registry identifies a mean and standard deviation for the primary continuous outcome, but does not describe how paired observations or possible dependence among mosquitoes within cups were modeled. Those design features can affect the interpretation and precision of an assay comparison.
Statistically, the trial matters because it asks a measurement-performance question in a defined repeated-assay setting. The registry record supplies context for that question, but numerical findings and methodological details are necessary before drawing conclusions about the degree of consistency.
14. Limitations
- The posted summary statistics are the only data these results rest on; patient-level data, full protocols and statistical analysis plans are not available.
- Recalculated results are simple, unadjusted calculations from posted summaries. They do not replace the sponsor's prespecified analysis.
A central limitation of the registry record is that it does not report numerical results for the primary outcome. Without Day 1 and Day 2 estimates or an agreement analysis, the study’s stated reproducibility objective cannot be evaluated from the record. Confidence intervals and other measures of uncertainty are also not reported.
The analysis description is incomplete for assessing the measurement structure. The registry does not state the statistical unit, how repeated observations were paired, whether dependence within cups was addressed, or how gametocyte density was considered. It lists a mean and standard deviation for the continuous primary outcome, but does not provide details about distributional assumptions or model choice. The statistical analysis plan is not reported in the registry record.
The endpoint is restricted to surviving mosquitoes. The record notes that the number analyzed reflects survival after 9 days and that one participant without surviving mosquitoes in any assay was excluded. It does not provide survival counts by cup or time point, additional missingness details, or sensitivity analyses. This limits assessment of whether the observed mosquitoes represent the intended assay population.
The registry identifies one Study Volunteers group and states that allocation was not described; it is not a like-for-like control comparison. The intervention listing includes primaquine and artemether/lumefantrine, but the arm description is incomplete and does not explain the full treatment schedule. Consequently, treatment effects cannot be separated from the assay-repeatability question using this record.
Finally, the registry does not report a criterion defining acceptable reproducibility, a sample-size justification, interim analyses, or multiplicity procedures. These omissions limit interpretation of both the planned inferential framework and the strength of any conclusion that might be drawn from results outside the registry record.
15. Related Tutorials
16. Related Statistical Calculators
17. Sources
- ClinicalTrials.gov: Clinical Investigation Study to Evaluate the Consistency and Reproducibility of Two Consecutive Mosquito Feeding Assays — NCT04666350.
- Registry record last updated 2024-04-29; status last verified 2022-12.
- Results first posted 2024-04-29.
- Data retrieved from ClinicalTrials.gov on 2026-11-09.
- Group sizes, means, event counts and sponsor-reported p-values are reproduced as posted. Effect estimates, confidence intervals and p-values not attributed to the sponsor are independently calculated from the posted summary statistics.
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18. Record Summary
NCT04666350 is a completed clinical investigation sponsored by PATH, with enrollment of 42. The registry describes two direct skin feeding assays performed on Day 1 and Day 2 within a 24-hour interval in adults with varying Plasmodium falciparum gametocyte densities. The primary outcome is oocyst prevalence among surviving mosquitoes, assessed 9 days after feeding.
The primary analysis population includes participants with available data; one participant without surviving mosquitoes in any assay was excluded. The registry lists oocyst density, sporozoite prevalence, and sporozoite density as secondary outcomes. It reports 0 serious adverse events, 32 participants affected by other adverse events, and 0 deaths among the listed safety figures.
Numerical primary and secondary outcome results, the statistical analysis plan, detailed missing-data methods, and a prespecified reproducibility criterion are not reported in the registry record. Accordingly, the record describes the study question and outcome definitions but does not permit quantification of assay consistency.