Experimental analysis of the liquid-feeding mechanism of the butterfly Pieris rapae.

The butterfly Pieirs rapae drinks liquid using a long proboscis. A high pressure gradient is induced in the proboscis when cibarial pump muscles contract. However, liquid feeding through the long proboscis poses a disadvantage of high flow resistance. Hence, butterflies may possess special features to compensate for this disadvantage and succeed in foraging. The main objective of this study is to analyze the liquid-feeding mechanism of butterflies. The systaltic motion of the cibarial pump organ was visualized using the synchrotron X-ray imaging technique. In addition, an ellipsoidal pump model was established based on synchrotron X-ray micro-computed tomography. To determine the relationship between the cyclic variation of the pump volume and the liquid-feeding flow, velocity fields of the intake flow at the tip of the proboscis were measured using micro-particle image velocimetry. Reynolds and Womersley numbers of liquid-feeding flow in the proboscis were ~1.40 and 0.129, respectively. The liquid-feeding flow could be characterized as a quasi-steady state laminar flow. Considering these results, we analyzed the dimensions of the feeding apparatus on the basis of minimum energy consumption during the liquid-feeding process. The relationship between the proboscis and the cibarial pump was determined when minimum energy consumption occurs. As a result, the volume of the cibarial pump is proportional to the cube of the radius of the proboscis. It seems that the liquid-feeding system of butterflies and other long-proboscid insects follow the cube relationship. The present results provide insights into the feeding strategies of liquid-feeding butterflies.

Liquid-intake flow around the tip of butterfly proboscis.

Butterflies drink liquid through a slender proboscis using a large pressure gradient induced by the systaltic operation of a muscular pump inside their head. Although the proboscis is a naturally well-designed coiled micro conduit for liquid uptake and deployment, it has been regarded as a simple straw connected to the muscular pump. There are few studies on the transport of liquid food in the proboscis of a liquid-feeding butterfly. To understand the liquid-feeding mechanism in the proboscis of butterflies, the intake flow around the tip of the proboscis was investigated in detail. In this study, the intake flow was quantitatively visualized using a micro-PIV (particle image velocimetry) velocity field measurement technique. As a result, the liquid-feeding process consists of an intake phase, an ejection phase and a rest phase. When butterflies drink pooled liquid, the liquid is not sucked into the apical tip of the proboscis, but into the dorsal linkage aligned longitudinally along the proboscis. To analyze main characteristics of the intake flow around a butterfly proboscis, a theoretical model was established by assuming that liquid is sucked into a line sink whose suction rate linearly decreases proximally. In addition, the intake flow around the tip of a female mosquito׳s proboscis which has a distinct terminal opening was also visualized and modeled for comparison. The present results would be helpful to understand the liquid-feeding mechanism of a butterfly.

Detection of heparin in the salivary gland and midgut of Aedes togoi.

Mosquitoes secrete saliva that contains biological substances, including anticoagulants that counteract a host's hemostatic response and prevent blood clotting during blood feeding. This study aimed to detect heparin, an anticoagulant in Aedes togoi using an immunohistochemical detection method, in the salivary canal, salivary gland, and midgut of male and female mosquitoes. Comparisons showed that female mosquitoes contained higher concentrations of heparin than male mosquitoes. On average, the level of heparin was higher in blood-fed female mosquitoes than in non-blood-fed female mosquitoes. Heparin concentrations were higher in the midgut than in the salivary gland. This indicates presence of heparin in tissues of A. togoi.

Time-resolved X-ray PIV technique for diagnosing opaque biofluid flow with insufficient X-ray fluxes.

X-ray imaging is used to visualize the biofluid flow phenomena in a nondestructive manner. A technique currently used for quantitative visualization is X-ray particle image velocimetry (PIV). Although this technique provides a high spatial resolution (less than 10 µm), significant hemodynamic parameters are difficult to obtain under actual physiological conditions because of the limited temporal resolution of the technique, which in turn is due to the relatively long exposure time (~10 ms) involved in X-ray imaging. This study combines an image intensifier with a high-speed camera to reduce exposure time, thereby improving temporal resolution. The image intensifier amplifies light flux by emitting secondary electrons in the micro-channel plate. The increased incident light flux greatly reduces the exposure time (below 200 µs). The proposed X-ray PIV system was applied to high-speed blood flows in a tube, and the velocity field information was successfully obtained. The time-resolved X-ray PIV system can be employed to investigate blood flows at beamlines with insufficient X-ray fluxes under specific physiological conditions. This method facilitates understanding of the basic hemodynamic characteristics and pathological mechanism of cardiovascular diseases.

Effect of fluid viscosity on the liquid-feeding flow phenomena of a female mosquito.

Liquid-sucking phenomena by the two-pump system of female mosquitoes were investigated to understand the feeding mechanism. In most previous experimental studies on liquid-feeding insects, the net increase of mass was divided by the feeding time and fluid density to evaluate the intake rate. However, this weighting method is not so precise for mosquitoes, because they are too lightweight to measure the gain of mass accurately. In this study, the intake rate of female mosquitoes feeding on various sucrose solutions was estimated using a micro-particle image velocimetry technique. As the sucrose concentration increased from 1% to 50%, the intake rate decreased from 17.3 to 5.8 nl s(-1). In addition, the temporal volume variations of the two pump chambers were estimated based on the velocity and acceleration information of the flow at the center of the food canal of the proboscis. One pumping period was divided into four elementary phases, which are related to the different operational modes of the two pumps. According to the hypothetical model established in this study, the phase shift () between the two pump chambers increases from 14 to 28 ms and the percentage of reverse flow to forward flow in a pumping period decreases from 7.6% to 1.7% with increasing viscosity. The developed analytical methodology thus aids in the study of an insect's feeding mechanism.

Experimental analysis of the blood-sucking mechanism of female mosquitoes.

Pioneering studies have been conducted to reveal the functional characteristics of the two-pump system of the female mosquito. Mosquitoes are equipped with two pumping organs located in the head: the cibarial (CP) and the pharyngeal (PP) pumps. To analyze the functional relationship of these pumps during the blood-sucking process, micro-particle image velocimetry (PIV) and synchrotron X-ray micro-imaging were employed. The two pumps were found to be well coordinated with a phase shift (α) and time shift (ß) but to have distinct functions in the liquid-sucking process. The first pump (CP) starts to expand first, and then the second pump (PP) expands in advance with a time shift (ß) before the first pump (CP) begins to contract, playing a key role in improving pumping performance. The systaltic motion of the two pumps works systematically in a well-coordinated manner. In addition, the pumping performance of blood-sucking female mosquitoes is demonstrated to be superior to that of nectar-eating male mosquitoes. Intake flow rate is maximized by reducing the relaxation time of the CP and increasing the pumping frequency.

The performance of bioinspired valveless piezoelectric micropump with respect to viscosity change.

This study investigated the effect of the serial connection of two pumping chambers on transport of liquid with increased viscosity. A serially connected valveless piezoelectric micropump was fabricated inspired by the liquid-feeding strategy of a female mosquito drinking liquid with a wide range of viscosities, from nectar to blood. The performance of the micropump was investigated by varying the viscosity of working liquid. Results showed that the optimal phase difference between the two chambers was 180° out-of-phase for all viscosity conditions. The two chambers operating at 180° out-of-phase exhibited higher pumping performance compared with the sum of each single chamber solely actuated, when viscosity increased. The flow patterns in the micropump showed that the rectification efficiency improved with the increase in viscosity. Results indicated that the serially connected valveless piezoelectric micropump is more robust to the increase of viscosity than a single-chamber piezoelectric micropump. This study would be helpful in the design of microfluidic devices for transporting liquids with a wide range of viscosities.

Synchrotron X-ray microscopic computed tomography of the pump system of a female mosquito.

The pumping organ of blood-sucking female mosquitoes has a three-dimensional (3D) structure. However, conventional two-dimensional imaging methods are insufficient for visualizing the 3D structure in detail. Furthermore, their 3D imaging tasks are highly time consuming and sample preparation process requires elaborate skill. Among 3D imaging techniques, synchrotron X-ray microscopic computed tomography (SR-µCT) is especially suitable for small insects with opaque cuticles, such as mosquitoes. In this study, the 3D morphological structure of the pump system of a female mosquito was visualized using SR-µCT. Expandable volume capacities of two pump chambers were measured for several mosquito samples of similar size. To verify the cross-sectional images acquired by SR-µCT, complementary paraffin-sectioning data were compared.

Chitosan microparticles incorporating gold as an enhanced contrast flow tracer in dynamic X-ray imaging.

In situ monitoring of a biofluid can provide important information on circulatory disorders and a basic understanding on the metabolic mechanisms of living organisms. X-ray imaging has significant advantages as one of the most popular diagnostic tools to seethrough various biological systems. Particle traced velocity field measurement is one of the most popular methods for quantitative analysis of dynamic flow motion. In this study we have developed chitosan microparticles incorporating gold nanoparticles (AuNP) as a new enhanced contrast flow tracer for dynamic X-ray imaging. Gold is a useful material possessing high X-ray absorption ability and also biocompatibility. We chose chitosan as an AuNP delivery system because it can effectively trap AuNPs at high yield. In particular, the unique gold ion reduction ability of and compatibility with surface-modified chitosans are effectively utilized. The physical properties of the Au-chitosan microparticles can be controlled by varying the molecular weight of the chitosan employed and the AuNP incorporation methodology. The environment of the particles and the type of applied X-ray essentially determine the imaging efficiency. The designed chitosan microparticles incorporating Au have been successfully applied to track the digestive mechanisms occurring in delicate insects such as live mosquitoes.

Experimental study on the fluid mechanics of blood sucking in the proboscis of a female mosquito.

Female mosquitoes are known to have a magnificent micro-scale pumping system that can transport small quantities of blood very effectively. To understand the dynamic characteristics of blood flow inside female mosquitoes, the measurement technique that is capable of measuring instantaneous flow fields of a biological sample at micrometer scales is required. In this study, the blood-sucking flow inside a female mosquito's food canal was measured in vivo using a micro particle image velocimetry (micro-PIV) velocity field measurement technique with high-temporal resolution. The volumetric flow rate (Q) and the time-averaged feeding speed (V) based on the diameter of the food canal (D) was found to be 5.751 x 10(-3) mm3/s and 0.416 cm/s, respectively. Spectral analysis on the velocity waveform shows a clear peak at 6.1 Hz, indicating distinct pulsatile blood-sucking characteristics. The Womersley number (alpha) was about 0.117 and the velocity profile of the blood flow inside the proboscis has a parabolic Hagen-Poiseuille flow pattern when alpha is much smaller than 1.