Insecticidal roof barriers mounted on untreated bed nets can be as effective against Anopheles gambiae as regular insecticide-treated bed nets.

Barrier bednets (BBnets), regular bednets with a vertical insecticidal panel to target mosquitoes above the bednet roof, where they are most active, have the potential to improve existing Insecticidal Treated Bednets (ITNs), by reducing the quantity of insecticide required per net, reducing the toxic risks to those using the net, potentially increasing insecticide choice. We evaluated the performance of PermaNet 3.0 (P3) and untreated (Ut) bed nets with and without pyrethroid and piperonyl butoxide roof barriers in killing pyrethroid-resistant and susceptible Anopheles gambiae, simultaneously video-recording mosquito flight tracks. Bioassay results showed that treated roof barriers, particularly the longitudinal P3 barrier (P3L) could be an effective addition to a bed net: P3 + P3L were consistently significantly more effective than the reference P3 bednet while performance of untreated nets could be raised to equal that of the reference P3 following the addition of a P3 barrier. The BBnet's potential to augment existing bednets and enhance their performance is considered.

Impacts of dual active-ingredient bed nets on the behavioural responses of pyrethroid resistant Anopheles gambiae determined by room-scale infrared video tracking.

BACKGROUND: The success of insecticide treated bed nets (ITNs) for malaria vector control in Africa relies on the behaviour of various species of Anopheles. Previous research has described mosquito behavioural alterations resulting from widespread ITN coverage, which could result in a decrease in net efficacy. Here, behaviours were compared including timings of net contact, willingness to refeed and longevity post-exposure to two next-generation nets, PermaNet® 3.0 (P3 net) and Interceptor® G2 (IG2 net) in comparison with a standard pyrethroid-only net (Olyset Net™ (OL net)) and an untreated net. METHODS: Susceptible and resistant Anopheles gambiae mosquitoes were exposed to the nets with a human volunteer host in a room-scale assay. Mosquito movements were tracked for 2 h using an infrared video system, collecting flight trajectory, spatial position and net contact data. Post-assay, mosquitoes were monitored for a range of sublethal insecticide effects. RESULTS: Mosquito net contact was focused predominantly on the roof for all four bed nets. A steep decay in activity was observed for both susceptible strains when P3 net and OL net were present and with IG2 net for one of the two susceptible strains. Total mosquito activity was higher around untreated nets than ITNs. There was no difference in total activity, the number, or duration, of net contact, between any mosquito strain, with similar behaviours recorded in susceptible and resistant strains at all ITNs. OL net, P3 net and IG2 net all killed over 90% of susceptible mosquitoes 24 h after exposure, but this effect was not seen with resistant mosquitoes where mortality ranged from 16 to 72%. All treated nets reduced the willingness of resistant strains to re-feed when offered blood 1-h post-exposure, with a more pronounced effect seen with P3 net and OL net than IG2 net. CONCLUSION: These are the first results to provide an in-depth description of the behaviour of susceptible and resistant Anopheles gambiae strains around next-generation bed nets using a room-scale tracking system to capture multiple behaviours. These results indicate that there is no major difference in behavioural responses between mosquito strains of differing pyrethroid susceptibility when exposed to these new ITNs under the experimental conditions used.

Improved three-dimensional localization of multiple small objects in close proximity in digital holography.

Using intensity gradient- or sparsity-based focus metrics, the ability to accurately localize the three-dimensional (3D) position of a small object in a digital holographic reconstruction of a large field of view is hindered in the presence of multiple nearby objects. A more accurate alternative method for 3D localization, based on evaluation of the complex reconstructed volume, is proposed. Simulations and experimental data demonstrate a reduction in depth positional error for single objects and a notably improved axial resolution of multiple objects in close proximity.

Diffuse retro-reflective imaging for improved video tracking of mosquitoes at human baited bednets.

Robust imaging techniques for tracking insects have been essential tools in numerous laboratory and field studies on pests, beneficial insects and model systems. Recent innovations in optical imaging systems and associated signal processing have enabled detailed characterization of nocturnal mosquito behaviour around bednets and improvements in bednet design, a global essential for protecting populations against malaria. Nonetheless, there remain challenges around ease of use for large-scale in situ recordings and extracting data reliably in the critical areas of the bednet where the optical signal is attenuated. Here, we introduce a retro-reflective screen at the back of the measurement volume, which can simultaneously provide diffuse illumination, and remove optical alignment issues while requiring only one-sided access to the measurement space. The illumination becomes significantly more uniform, although noise removal algorithms are needed to reduce the effects of shot noise, particularly across low-intensity bednet regions. By systematically introducing mosquitoes in front of and behind the bednet in laboratory experiments, we are able to demonstrate robust tracking in these challenging areas. Overall, the retro-reflective imaging set-up delivers mosquito segmentation rates in excess of 90% compared to less than 70% with backlit systems.

Barrier bednets target malaria vectors and expand the range of usable insecticides.

Transmission of Plasmodium falciparum malaria parasites occurs when nocturnal Anopheles mosquito vectors feed on human blood. In Africa, where malaria burden is highest, bednets treated with pyrethroid insecticide were highly effective in preventing mosquito bites and reducing transmission, and essential to achieving unprecedented reductions in malaria until 2015 (ref. 1). Since then, progress has stalled2, and with insecticidal bednets losing efficacy against pyrethroid-resistant Anopheles vectors3,4, methods that restore performance are urgently needed to eliminate any risk of malaria returning to the levels seen before their widespread use throughout sub-Saharan Africa5. Here, we show that the primary malaria vector Anopheles gambiae is targeted and killed by small insecticidal net barriers positioned above a standard bednet in a spatial region of high mosquito activity but zero contact with sleepers, opening the way for deploying many more insecticides on bednets than is currently possible. Tested against wild pyrethroid-resistant A. gambiae in Burkina Faso, pyrethroid bednets with organophosphate barriers achieved significantly higher killing rates than bednets alone. Treated barriers on untreated bednets were equally effective, without significant loss of personal protection. Mathematical modelling of transmission dynamics predicted reductions in clinical malaria incidence with barrier bednets that matched those of 'next-generation' nets recommended by the World Health Organization against resistant vectors. Mathematical models of mosquito-barrier interactions identified alternative barrier designs to increase performance. Barrier bednets that overcome insecticide resistance are feasible using existing insecticides and production technology, and early implementation of affordable vector control tools is a realistic prospect.

Host-seeking activity of a Tanzanian population of Anopheles arabiensis at an insecticide treated bed net.

BACKGROUND: Understanding how mosquitoes respond to long lasting insecticide treated nets (LLINs) is fundamental to sustaining the effectiveness of this essential control tool. We report on studies with a tracking system to investigate behaviour of wild anophelines at an LLIN, in an experimental hut at a rural site in Mwanza, Tanzania. METHODS: Groups of adult female mosquitoes (n = 10 per replicate) reared from larvae of a local population, identified as predominantly (95%) Anopheles arabiensis, were released in the hut. An infrared video tracking system recorded flight and net contact activity over 1 h as the mosquitoes attempted to reach a supine human volunteer within a bed net (either a deltamethrin-treated LLIN or an untreated control net). A range of activities, including flight path, position in relation to the bed net and duration of net contact, were quantified and compared between treatments. RESULTS: The total time that female An. arabiensis spent in flight around LLINs was significantly lower than at untreated nets [F(1,10) = 9.26, p = 0.012], primarily due to a substantial reduction in the time mosquitoes spent in persistent 'bouncing' flight [F(1,10) = 18.48, p = 0.002]. Most activity occurred at the net roof but significantly less so with LLINs (56.8% of total) than untreated nets [85.0%; Χ2 (15) = 234.69, p < 0.001]. Activity levels at the bed net directly above the host torso were significantly higher with untreated nets (74.2%) than LLINs [38.4%; Χ2 (15) = 33.54, p = 0.004]. 'Visiting' and 'bouncing' rates were highest above the volunteer's chest in untreated nets (39.9 and 50.4%, respectively) and LLINs [29.9 and 42.4%; Χ2 (13) = 89.91, p < 0.001; Χ2 (9) = 45.73, p < 0.001]. Highest resting rates were above the torso in untreated nets [77%; Χ2 (9) = 63.12, p < 0.001], but in LLINs only 33.2% of resting occurred here [Χ2 (9) = 27.59, p = 0.001], with resting times spread between the short vertical side of the net adjacent to the volunteer's head (21.8%) and feet (16.2%). Duration of net contact by a single mosquito was estimated at 204-290 s on untreated nets and 46-82 s on LLINs. While latency to net contact was similar in both treatments, the reduction in activity over 60 min was significantly more rapid for LLINs [F(1,10) = 6.81, p = 0.026], reiterating an 'attract and kill' rather than a repellent mode of action. CONCLUSIONS: The study has demonstrated the potential for detailed investigations of behaviour of wild mosquito populations under field conditions. The results validate the findings of earlier laboratory studies on mosquito activity at LLINs, and reinforce the key role of multiple brief contacts at the net roof as the critical LLIN mode of action.

Infrared video tracking of Anopheles gambiae at insecticide-treated bed nets reveals rapid decisive impact after brief localised net contact.

Long-lasting insecticidal bed nets (LLINs) protect humans from malaria transmission and are fundamental to malaria control worldwide, but little is known of how mosquitoes interact with nets. Elucidating LLIN mode of action is essential to maintain or improve efficacy, an urgent need as emerging insecticide resistance threatens their future. Tracking multiple free-flying Anopheles gambiae responding to human-occupied bed nets in a novel large-scale system, we characterised key behaviours and events. Four behavioural modes with different levels of net contact were defined: swooping, visiting, bouncing and resting. Approximately 75% of all activity occurred at the bed net roof where multiple brief contacts were focussed above the occupant's torso. Total flight and net contact times were lower at LLINs than untreated nets but the essential character of the response was unaltered. LLINs did not repel mosquitoes but impacted rapidly: LLIN contact of less than 1 minute per mosquito during the first ten minutes reduced subsequent activity; after thirty minutes, activity at LLINs was negligible. Velocity measurements showed that mosquitoes detected nets, including unbaited untreated nets, prior to contact. This is the most complete characterisation of mosquito-LLIN interactions to date, and reveals many aspects of LLIN mode of action, important for developing the next generation of LLINs.

Algebraic solution for phase unwrapping problems in multiwavelength interferometry.

Recent advances in multiwavelength interferometry techniques [Appl. Opt.52, 5758 (2013)] give new insights to phase unwrapping problems and allow the fringe order information contained in the measured phase to be extracted with low computational effort. This work introduces an algebraic solution to the phase unwrapping problem that allows the direct calculation of the unknown integer fringe order. The procedure resembles beat-wavelength approaches, but provides greater flexibility in choosing the measurement wavelengths, a larger measurement range, and a higher robustness against noise, due to the ability to correct for errors during the calculation.

Method of excess fractions with application to absolute distance metrology: analytical solution.

Multiwavelength interferometry provides a solution to a number of applications in metrology for the measurement of optical path differences longer than the source wavelength. To this day, the method of excess fractions (EF) has proved to provide very long, unambiguous measurement ranges with the highest reliability for a given set of wavelengths and level of phase noise. This is achieved because EF combines the individual phase values in an equivalent least-square problem and evaluates the correspondence for all possible solutions. However, this procedure can be slow for a number of applications. In this paper, an analytical solution for EF is presented that allows the direct calculation of the unknown integer fringe order. It is shown that this solution is consistent with the other phase unwrapping approaches as beat wavelength or Chinese remainder theorem-based solutions, but moreover, it can be understood as a unified representation and solution of the fringe order problem.

Method of excess fractions with application to absolute distance metrology: wavelength selection and the effects of common error sources.

Multiwavelength interferometry (MWI) is a well established technique in the field of optical metrology. Previously, we have reported a theoretical analysis of the method of excess fractions that describes the mutual dependence of unambiguous measurement range, reliability, and the measurement wavelengths. In this paper wavelength, selection strategies are introduced that are built on the theoretical description and maximize the reliability in the calculated fringe order for a given measurement range, number of wavelengths, and level of phase noise. Practical implementation issues for an MWI interferometer are analyzed theoretically. It is shown that dispersion compensation is best implemented by use of reference measurements around absolute zero in the interferometer. Furthermore, the effects of wavelength uncertainty allow the ultimate performance of an MWI interferometer to be estimated.

Method of excess fractions with application to absolute distance metrology: theoretical analysis.

The method of excess fractions (EF) is well established to resolve the fringe order ambiguity generated in interferometric detection. Despite this background, multiwavelength interferometric absolute long distance measurements have only been reported with varying degrees of success. In this paper we present a theoretical model that can predict the unambiguous measurement range in EF based on the selected measurement wavelengths and phase noise. It is shown that beat wavelength solutions are a subset of this theoretical model. The performance of EF, for a given phase noise, is shown to be equivalent to beat techniques but offers many alternative sets of measurement wavelengths and therefore EF offer significantly greater flexibility in experimental design.

Absolute metrology by phase and frequency modulation for multiwavelength interferometry.

A fiber interferometer for absolute distance measurements is presented whereby wavelength variation is achieved via a sinusoidal strain modulation of a fiber Bragg grating to generate a series of beat wavelengths. The interferometer employs fiber laser sources where the design is based on the use of narrow-bandwidth fiber Bragg gratings. The accuracy of the beat wavelengths is improved compared to the use of multiple wavelengths measured with conventional optical spectrum analyzers or available wavemeters. Initial measurements are presented for beat wavelengths of 254.74 mm and 27.4 m over an optical path difference of 200 mm and 3.8 m, respectively. Combined with a two (or three) wavelength interferometer, this technique has the potential for ultrahigh dynamic range metrology ranging over several meters while preserving subfringe resolution and a low system complexity.

Simple calibration of a phase-based 3D imaging system based on uneven fringe projection.

Phase-based fringe projection metrology systems have been widely used to obtain the shape of 3D objects. One vital step is calibration, which defines the relationship between the phase and depth data. Existing calibration methods are complicated because of the dependence of the relationship on the pixel position. In this Letter, a simple calibration procedure is introduced based on an uneven fringe projection technique, in which the relationship between phase and depth becomes independent of the pixel position and can be represented by a single polynomial function for all pixels. Therefore, given a set of discrete points with a known phase and depth in the measuring volume, the coefficient set of the polynomial function can be determined. A white plate having discrete markers with known separation is used to calibrate the 3D imaging system. Experimental results demonstrate that the proposed calibration method is simple to apply and can build up an accurate relationship between phase and depth data.

Nanometric depth resolution from multi-focal images in microscopy.

We describe a method for tracking the position of small features in three dimensions from images recorded on a standard microscope with an inexpensive attachment between the microscope and the camera. The depth-measurement accuracy of this method is tested experimentally on a wide-field, inverted microscope and is shown to give approximately 8 nm depth resolution, over a specimen depth of approximately 6 µm, when using a 12-bit charge-coupled device (CCD) camera and very bright but unresolved particles. To assess low-flux limitations a theoretical model is used to derive an analytical expression for the minimum variance bound. The approximations used in the analytical treatment are tested using numerical simulations. It is concluded that approximately 14 nm depth resolution is achievable with flux levels available when tracking fluorescent sources in three dimensions in live-cell biology and that the method is suitable for three-dimensional photo-activated localization microscopy resolution. Sub-nanometre resolution could be achieved with photon-counting techniques at high flux levels.

Multiwavelength interferometry: extended range metrology.

We present an optimized method for multiwavelength interferometry that allows measurements beyond the largest beat wavelength. The approach exploits wavelength coincidence between two beat wavelengths in order to measure unambiguously over an extended range. Performance of the approach has been validated both through simulations and experimentally by means of a fiber interferometer for four measurement wavelengths. Initial results have demonstrated 1/200th of a fringe phase resolution, giving absolute metrology over 18.16 mm, or a dynamic range of 1 part in 2.4x10(6). With improved phase resolution the method has the potential to range over >100 m using femtosecond laser frequency comb sources.

Three-dimensional particle imaging by defocusing method with an annular aperture.

We propose an annular-aperture-based defocusing technique for three-dimensional (3D) particle metrology from a single camera view. This simple configuration has high optical efficiency and the ability to deal with overlapped defocused images. Initial results show that an uncertainty in depth of 23 microm can be achieved over a range of 10 mm for macroscopic systems. This method can also be applied in microscopy for the measurement of fluorescently doped microparticles, thus providing a promising solution for 3D flow metrology at both macroscales and microscales.

Uneven fringe projection for efficient calibration in high-resolution 3D shape metrology.

A novel uneven fringe projection technique is presented whereby nonuniformly spaced fringes are generated at a digital video projector to give evenly spaced fringes in the measurement volume. The proposed technique simplifies the relation between the measured phase and the object's depth independent of pixel position. This method needs just one coefficient set for calibration and depth calculation. With uneven fringe projection the shape data are referenced to a virtual plane instead of a physical reference plane, so an improved measurement with lower uncertainty is achieved. Further, the method can be combined with a radial lens distortion model. The theoretical foundation of the method is presented and experimentally validated to demonstrate the advantages of the uneven fringe projection approach compared with existing methods. Measurement results on a National Physical Laboratory (UK) "step standard" confirm the measurement uncertainty using the proposed method.

Three-dimensional particle imaging by wavefront sensing.

We present two methods for three-dimensional particle metrology from a single two-dimensional view. The techniques are based on wavefront sensing where the three-dimensional location of a particle is encoded into a single image plane. The first technique is based on multiplanar imaging, and the second produces three-dimensional location information via anamorphic distortion of the recorded images. Preliminary results show that an uncertainty of 8 microm in depth can be obtained for low-particle density over a thin plane, and an uncertainty of 30 microm for higher particle density over a 10 mm deep volume.

Time efficient color fringe projection system for 3D shape and color using optimum 3-frequency Selection.

We present a novel color fringe projection system to obtain absolute 3D shape and color of objects simultaneously. Optimum 3-frequency interferometry is used to produce time efficient analysis of the projected fringes by encoding three fringe sets of different pitch into the primary colors of a digital light projector and recording the information on a 3-chip color CCD camera. Phase shifting analysis is used to retrieve subwavelength phase information. Absolute phase across the field is calculated using the 3-frequency method independently at each pixel. Concurrent color data is also captured via the RGB channels of the CCD. Thus full-field absolute shape (XYZ) and color (RGB) can be obtained. In this paper we present the basis of the technique and preliminary results having addressed the issue of crosstalk between the color channels.

Fiber interferometer for simultaneous multiwavelength phase measurement with a broadband femtosecond laser.

We present a fiber interferometer for the simultaneous measurement of phase at multiple wavelengths from a single broadband femtosecond laser. Narrow-bandwidth fiber Bragg gratings isolate a particular frequency from the broad-bandwidth laser pulse produced. The multiwavelength phase data permit the unambiguous measurement range to be significantly increased compared with the wavelengths used in the interferometer. Preliminary experimental results are presented for a two-frequency sensor with an absolute range of 0.13 mm and associated dynamic range of 43,000:1.