Distinguishing the Cellular Transport of Folic Acid Conjugated Nano-Drugs among Different Cell Lines by Using Force Tracing Technique.

Folic acid (FA) is a ligand that has been renowned for its strong binding to FA receptor (FR), and the robustness of the specific interaction has led to the generation of multitudinous tumor-targeted nano-drug delivery systems. However, selecting the appropriate FA targeted nano-drugs according to types of cancerous cells to achieve a high effect is critical. Understanding of how the drug is transported through the cell membrane and is delivered intracellularly is very important in screening ideal targeted nano-drugs for cancerous changes in different organs. Herein, by using a force tracing technique based on atomic force microscopy (AFM), the dynamic process of FA-polyamidoamine-Doxorubicin (FA-PAMAM-DOX) entry into different tumor cells (HeLa and A549) and normal cells (Vero) was monitored in real time. The cell membrane transport efficacy of FA-PAMAM-DOX in tumor cells with an FR high overexpression level (HeLa) and FR low overexpression level (A549) is analyzed, which is significantly higher than that in normal cells (Vero), especially for HeLa cells. Subsequently, the intracellular delivery efficiency of FA-PAMAM-DOX in different cell lines was measured by using fluorescence imaging and AFM-based nanoindentation techniques. This report will help to discover the cellular transport mechanism of nano-drugs and screen out optimal therapeutic nano-drugs for different types of tumors.

Investigating the trans-membrane transport of HAIYPRH peptide-decorated nano-drugs.

Transferrin (Tf) has been effectively used to promote the cellular uptake of HAIYPRH (T7) peptide-conjugated nano-drugs. In this study, the enhancing effect of Tf on T7-decorated nano-drug transport was investigated using force tracing and nano-indentation techniques at a single-particle/cell level. Furthermore, the results were confirmed by ensemble fluorescence imaging.

Infantile Cranial Fasciitis: A Clinicopathologic Evaluation.

OBJECTIVE: To investigate the clinicopathologic features, immunophenotype, molecular genetic changes, and differential diagnosis of cranial fasciitis (CF). METHODS: The clinical manifestations, imaging, surgical technique, pathologic characteristics, special staining, and immunophenotype, as well as break-apart fluorescence in situ hybridization assay for USP6 of 19 CF cases were analyzed, retrospectively. RESULTS: The patients were 11 boys and 8 girls, aged 5 to 144 months, with a median age of 29 months. There were 5 cases (26.31%) in the temporal bone, 4 cases (21.05%) in the parietal bone, 3 cases (15.78%) in the occipital bone, 3 cases (15.78%) in the frontotemporal bone, 2 cases (10.52%) in the frontal bone, 1 case (5.26%) in the mastoid of middle ear, and 1 case (5.26%) in the external auditory canal. The main clinical manifestations were painless, with the presentation of masses that grew rapidly and frequently eroded the skull. There was no recurrence and no metastasis after the operation. Histologically, the lesion consists of spindle fibroblasts/myofibroblasts arranged in bundles, braided or atypical spokes. Mitotic figures could be seen, but not atypical forms. Immunohistochemical studies showed diffuse strong positive SMA and Vimentin in all CFs. These cells were negative for Calponin, Desmin, ß-catenin, S-100, and CD34. The ki-67 proliferation index was 5% to 10%. Ocin blue-PH2.5 staining showed blue-stained mucinous features in the stroma. The positive rate of USP6 gene rearrangement detected by fluorescence in situ hybridization assay was about 10.52%, and the positive rate was not related to age. All patients were observed for 2 to 124 months and showed no signs of recurrence or metastasis. CONCLUSIONS: In summary, CF was a benign pseudosarcomatous fasciitis that occurs in the skull of infants. Preoperative diagnosis and differential diagnosis were difficult. Computed tomography typing might be beneficial for imaging diagnosis, and pathologic examination might be the most reliable way to diagnose CF.

Clinicopathological Analysis of Sturge-Weber Syndrome with Focal Cortical Dysplasia FCD IIIc.

Objective: To investigate the clinicopathological features of children with Sturge-Weber syndrome and to analyze the correlation between the distribution area of leptomeningeal angiomatosis, the degree of cerebral cortical calcification, and the degree of cerebral atrophy associated with epileptic seizures. Methods: 10 children were diagnosed with SWS with FCD IIIc by histopathology and immunohistochemistry. Spearman correlation analysis was used to calculate the association of SWS with FCD IIIc and seizures in children. Results: The leptomeningeal angiomatosis area was markedly positively correlated with the degree of brain atrophy in 10 children with SWS (r = 0.783, p = 0.007). The distribution of leptomeningeal hemangiomatosis, the degree of cortical calcification, and brain atrophy were not significantly correlated with epilepsy. Conclusion: SWS may be accompanied by FCD IIIc. The more extensive the cerebral lobes of leptomeningeal angiomatosis in SWS, the more pronounced the brain atrophy.

The catalytic effect of RuM-C catalyst attached to carbon- based support for hydrogen evolution reaction.

The potential of converting traditional biomass into low-cost HER catalysts has broad application prospects. In this paper, fungus is used as a carbon-based carrier. The bimetallic catalyst RuM-C (M = V, Mo, W, Zn, Cu) was synthesized under inert gas protection at high temperature. The order of electrocatalytic activity is RuV-C > RuZn-C > RuW-C > RuMo-C > Ru-C > RuCu-C > BF-C, which indicates that RuV-C exhibits excellent HER activity. Due to its irregular sheet structure, the specific surface area of the catalyst is increased. Impressively, it exhibits extremely high catalytic activity for HER in 1 M KOH due to favorable kinetics and excellent specific activity. Consequently, the prepared RuV-C exhibited excellent and stable HER activity compared Ru-C with a low overpotential of 65.78 mV at the current densities of 10 mA cm-2and Tafel slope of 45.26 mV dec-1. The potential only decreased by 88 mV after 24 h of continuous testing, which indicates that the catalyst has outstanding stability. This work will provide positive inspiration for the promotion of a new Ru-based biomass HER electrocatalyst.

Assemblages of Plasmodium and Related Parasites in Birds with Different Migration Statuses.

Migratory birds spend several months in their breeding grounds in sympatry with local resident birds and relatively shorter periods of time at stopover sites. During migration, parasites may be transmitted between migratory and resident birds. However, to what extent they share these parasites remains unclear. In this study, we compared the assemblages of haemosporidian parasites in migratory, resident, and passing birds, as well as the correlations between parasite assemblages and host phylogeny. Compared with passing birds, migratory birds were more likely to share parasites with resident birds. Shared lineages showed significantly higher prevalence rates than other lineages, indicating that common parasites are more likely to spill over from the current host to other birds. For shared lineages, the prevalence was significantly higher in resident birds than in migratory birds, suggesting that migratory birds pick up parasites at their breeding ground. Among the shared lineages, almost two-thirds presented no phylogenetic signal in their prevalence, indicating that parasite transmission among host species is weakly or not correlated with host phylogeny. Moreover, similarities between parasite assemblages are not correlated with either migration status or the phylogeny of hosts. Our results show that the prevalence, rather than host phylogeny, plays a central role in parasite transmission between migratory and resident birds in breeding grounds.

Revealing the Cell Entry Dynamic Mechanism of Single Rabies Virus Particle.

The rabies virus is a neurotropic virus that causes fatal diseases in humans and animals. Although studying the interactions between a single rabies virus and the cell membrane is necessary for understanding the pathogenesis, the internalization dynamic mechanism of single rabies virus in living cells remains largely elusive. Here, we utilized a novel force tracing technique based on atomic force microscopy(AFM) to record the process of single viral entry into host cell. We revealed that the force of the rabies virus internalization distributed at (65±25) pN, and the time was identified by two peaks with spacings of (237.2±59.1) and (790.3±134.4) ms with the corresponding speed of 0.12 and 0.04 µm/s, respectively. Our results provide insight into the effects of viral shape during the endocytosis process. This report will be meaningful for understanding the dynamic mechanism of rabies virus early infection. Electronic Supplementary Material: Supplementary material is available in the online version of this article at 10.1007/s40242-022-2069-y.

Revealing the Dynamic Mechanism by Which Transferrin Promotes the Cellular Uptake of HAIYPRH Peptide-Conjugated Nanostructures by Force Tracing.

The HAIYPRH (T7) peptide has been widely used as a ligand for constructing tumor-targeted nanodrug delivery systems since it can target the transferrin receptor (TfR) and then enter cells easily with the help of transferrin (Tf). However, the dynamic mechanism by which transferrin promotes the entry of T7-conjugated nanostructures into cells remains unclear. Herein, a force tracing technique based on atomic force microscopy (AFM) was used to track the ultrafast dynamic process of a T7-conjugated gold nanoparticle (AuNP-T7) entering a cell at the single-particle level in real time. Tf helped decrease the endocytosis force and increase the endocytosis speed of AuNP-T7 in A549 cells. However, Tf only increased the endocytosis speed of AuNP-T7 in HeLa cells. In contrast, in Vero cells without TfR overexpression, Tf decreased the endocytosis speed. This report provides important insights for redesigning and developing T7-conjugated nanodrug carriers in targeted nanodrug delivery systems.

Highly sensitive detection of DNA damage in living cells by SERS and electrochemical measurements using a flexible gold nanoelectrode.

Guanine (G) oxidation products, such as 8-hydroxy-2'-deoxyguanosine (8-OHdG) and 8-oxo-guanine (8-OXOG), have been widely studied as promising biomarkers for DNA oxidative damage. In this work, we develop a new method to detect G oxidative products released from live cells after chromium (vi) ion or hydrogen peroxide treatments by using a glass nanopipette-based flexible gold nanoelectrode (fGNE). Specific response to G oxidative products with high sensitivity can be detected from the fGNE tip through integrated electrochemical measurements and surface-enhanced Raman spectroscopy. The fGNE apex can be positioned very close to the cell membrane noninvasively because of its high flexibility and nanoscale tip size. With the assistance of the electrophoretic force, the fGNEs can effectively collect and detect the G-derived DNA damage products released from individual cells in the cell culture medium with high sensitivity.

Improvement of Sensing Properties for Copper Phthalocyanine Sensors Based on Polymer Nanofibers Scaffolds.

An effectual and understandable route for the fabrication techniques of stereoscopic NO2 sensor is provided in this work. As the gas-sensing layer of the sensor, copper phthalocyanine (CuPc) grew on the top of poly(vinyl alcohol) (PVA) nanofibers (NFs). The sensitivity of the CuPc/PVA NFs stereoscopic sensors to NO2 was over 829%/ppm, while the sensitivity of the continuous CuPc films sensors was 2 orders of magnitude lower than that of the stereoscopic ones. To the responsivities at 25 ppm of NO2, the CuPc/PVA NFs stereoscopic sensors were about four times stronger than that of the continuous CuPc films sensors. For the recovery time, the CuPc/PVA NFs stereoscopic sensors were over eight times faster than the continuous CuPc films sensors. This general tactic can be used to prepare various toxic gas sensors to improve the overall performance of the devices.

High proton conduction at above 100 °C mediated by hydrogen bonding in a lanthanide metal-organic framework.

A lanthanide metal-organic framework (MOF) compound of the formulation [Eu2(CO3)(ox)2(H2O)2]·4H2O (1, ox = oxalate) was prepared by hydrothermal synthesis with its structure determined crystallographically. Temperature-dependent but humidity-independent high proton conduction was observed with a maximum of 2.08 × 10(-3) S cm(-1) achieved at 150 °C, well above the normal boiling point of water. Results from detailed structural analyses, comparative measurements of conductivities using regular and deuterated samples, anisotropic conductivity measurements using a single-crystal sample, and variable-temperature photoluminescence studies collectively establish that the protons furnished by the Eu(III)-bound and activated aqua ligands are the charge carriers and that the transport of proton is mediated along the crystallographic a-axis by ordered hydrogen-bonded arrays involving both aqua ligands and adjacent oxalate groups in the channels of the open framework. Proton conduction was enhanced with the increase of temperature from room temperature to about 150 °C, which can be rationalized in terms of thermal activation of the aqua ligands and the facilitated transport between aqua and adjacent oxalate ligands. A complete thermal loss of the aqua ligands occurred at about 160 °C, resulting in the disintegration of the hydrogen-bonded pathway for proton transport and a precipitous drop in conductivity. However, the structural integrity of the MOF was maintained up to 350 °C, and upon rehydration, the original structure with the hydrogen-bonded arrays was restored, and so was its high proton-conduction ability.

Methylene blue modulates huntingtin aggregation intermediates and is protective in Huntington's disease models.

Huntington's disease (HD) is a devastating neurodegenerative disorder with no disease-modifying treatments available. The disease is caused by expansion of a CAG trinucleotide repeat and manifests with progressive motor abnormalities, psychiatric symptoms, and cognitive decline. Expression of an expanded polyglutamine repeat within the Huntingtin (Htt) protein impacts numerous cellular processes, including protein folding and clearance. A hallmark of the disease is the progressive formation of inclusions that represent the culmination of a complex aggregation process. Methylene blue (MB), has been shown to modulate aggregation of amyloidogenic disease proteins. We investigated whether MB could impact mutant Htt-mediated aggregation and neurotoxicity. MB inhibited recombinant protein aggregation in vitro, even when added to preformed oligomers and fibrils. MB also decreased oligomer number and size and decreased accumulation of insoluble mutant Htt in cells. In functional assays, MB increased survival of primary cortical neurons transduced with mutant Htt, reduced neurodegeneration and aggregation in a Drosophila melanogaster model of HD, and reduced disease phenotypes in R6/2 HD modeled mice. Furthermore, MB treatment also promoted an increase in levels of BDNF RNA and protein in vivo. Thus, MB, which is well tolerated and used in humans, has therapeutic potential for HD.

Identification of novel potentially toxic oligomers formed in vitro from mammalian-derived expanded huntingtin exon-1 protein.

Huntington disease is a genetic neurodegenerative disorder that arises from an expanded polyglutamine region in the N terminus of the HD gene product, huntingtin. Protein inclusions comprised of N-terminal fragments of mutant huntingtin are a characteristic feature of disease, though are likely to play a protective role rather than a causative one in neurodegeneration. Soluble oligomeric assemblies of huntingtin formed early in the aggregation process are candidate toxic species in HD. In the present study, we established an in vitro system to generate recombinant huntingtin in mammalian cells. Using both denaturing and native gel analysis, we have identified novel oligomeric forms of mammalian-derived expanded huntingtin exon-1 N-terminal fragment. These species are transient and were not previously detected using bacterially expressed exon-1 protein. Importantly, these species are recognized by 3B5H10, an antibody that recognizes a two-stranded hairpin conformation of expanded polyglutamine believed to be associated with a toxic form of huntingtin. Interestingly, comparable oligomeric species were not observed for expanded huntingtin shortstop, a 117-amino acid fragment of huntingtin shown previously in mammalian cell lines and transgenic mice, and here in primary cortical neurons, to be non-toxic. Further, we demonstrate that expanded huntingtin shortstop has a reduced ability to form amyloid-like fibrils characteristic of the aggregation pathway for toxic expanded polyglutamine proteins. Taken together, these data provide a possible candidate toxic species in HD. In addition, these studies demonstrate the fundamental differences in early aggregation events between mutant huntingtin exon-1 and shortstop proteins that may underlie the differences in toxicity.

Four new three-dimensional polyoxometalate-based metal-organic frameworks constructed from [Mo6O18(O3AsPh)2]4- polyoxoanions and copper(I)-organic fragments: syntheses, structures, electrochemistry, and photocatalysis properties.

Four novel polyoxometalate-based copper(I)-organic frameworks, namely, [Cu(I)(2)(cis-L1)(2)][Cu(I)(2)(trans-L1)(2)Mo(6)O(18)(O(3)AsPh)(2)] (1), [Cu(I)(4)(L2)(4)Mo(6)O(18)(O(3)AsPh)(2)] (2), [Cu(I)(4)(L3)(4)Mo(6)O(18)(O(3)AsPh)(2)] (3), and [Cu(I)(4)(L4)(2)Mo(6)O(18)(O(3)AsPh)(2)] (4) (L1 = 1,3-bis(1,2,4-triazol-1-yl)propane, L2 = 1,4-bis(1,2,4-triazol-1-yl)butane, L3 = 1,5-bis(1,2,4-triazol-1-yl)pentane, and L4 = 1,6-bis(1,2,4-triazol-1-yl)hexane), have been successfully synthesized under hydrothermal conditions. Their structures have been determined by single-crystal X-ray diffraction analyses and further characterized by elemental analyses, infrared spectra (IR), UV-vis spectra, powder X-ray diffraction (PXRD), and thermogravimetric (TG) analyses. Compound 1 is composed of two crystallographically independent and distinct polymeric motifs: one-dimensional (1D) S-shaped chain and two-dimensional (2D) undulated layer. The S-shaped chains penetrated into the 2D parallel layers to generate an unusual 1D + 2D → three-dimensional (3D) polypseudo-rotaxane framework. In 2, the As(2)Mo(6) polyoxoanions in tetradentate modes link four neighboring -L2-Cu-L2- chains to produce a rare 3D trinodal (3,4)-connected self-penetrated framework with Point Symbol of (8(3))(2)(8(2)·12(4)). In 3, adjacent Cu(I) atoms are linked by As(2)Mo(6) polyoxoanions and L3 ligands into a 2D layer. The layers are further interlocked by the two nearest neighboring ones to form a 3D polycatenated framework. In 4, L4 ligands bridge four Cu(I) atoms to yield 2D wavelike layers, which are further linked by the octadentate As(2)Mo(6) polyoxoanions to form a 3D tetranodal (3,4,6)-connected framework with Point Symbol of (6(3))(4·6(3)·8(2))(6(4)·8(2))(4(2)·6(2)·8(10)·10). In addition, the photocatalytic activities of compounds 1, 3, and 4 for decomposition of methylene blue (MB) under UV light have been investigated. Moreover, their electrochemical properties have also been studied in 1 M H(2)SO(4) aqueous solution.

Disrupted functional networks within white-matter served as neural features in adolescent patients with conduct disorder.

BACKGROUND: Conduct disorder (CD) has been conceptualized as a psychiatric disorder associated with white-matter (WM) structural abnormalities. Although diffusion tensor imaging could identify WM structural architecture changes, it cannot characterize functional connectivity (FC) within WM. Few studies have focused on disentangling the WM dysfunctions in CD patients by using functional magnetic resonance imaging (fMRI). METHODS: The resting-state fMRI data were first obtained from both adolescent CD and typically developing (TD) controls. A voxel-based clustering analysis was utilized to identify the large-scale WM FC networks. Then, we examined the disrupted WM network features in CD, and further investigated whether these features could predict the impulsive symptoms in CD using support vector regression prediction model. RESULTS: We identified 11 WM functional networks. Compared with TDs, CD patients showed increased FCs between occipital network (ON) and superior temporal network (STN), between orbitofrontal network (OFN) and corona radiate network (CRN), as well as between deep network and CRN. Further, the disrupted FCs between ON and STN and between OFN and CRN were significantly negatively associated with non-planning impulsivity scores in CD. Moreover, the disrupted WM networks could be served as features to predict the motor impulsivity scores in CD. CONCLUSIONS: Our results provided further support on the existence of WM functional networks and could extended our knowledge about the WM functional abnormalities related with emotional and perception processing in CD patients from the view of WM dysfunction.

Multimodal Brain Image Fusion Based on Improved Rolling Guidance Filter and Wiener Filter.

Medical image fusion technology can integrate complementary information from different modality medical images to provide a more complete and accurate description of the specific diagnosed object, which is very helpful for image-guided clinical diagnosis and treatment. This paper proposes an effective brain image fusion framework based on improved rolling guidance filter (IRGF). Firstly, input images are decomposed into base layers and detail layers using the IRGF and Wiener filter. Secondly, the visual saliency maps of the input image are computed by pixel-level saliency value, and the weight maps of detail layers are constructed by max-absolute strategy and are further smoothed with Gaussian filter, the purpose of which is to make the fused image appear more naturally and more suitable for human visual perception. Lastly, base layers are fused by visual saliency map based fusion rule and the corresponding weight maps from detail layers are fused by the weighted least squares optimization scheme. Experimental results testify that our method is superior to some state-of-the-art methods in both subjective and objective assessments.

High-strength and anti-biofouling nanofiber membranes for enhanced uranium recovery from seawater and wastewater.

Ultrathin fibers can increase the contact area between adsorbents and seawater during the uranium extraction process; however, their construction usually aggravates the complex spinning technology and lowers their mechanical strength. Meanwhile, high strength and antifouling ability are essential for ocean adsorbents to withstand the complex natural environment and microbial systems. Herein, we design high-strength and anti-biofouling poly(amidoxime) nanofiber membranes (HA-PAO NFMs) via a supramolecular crosslinking. Bacterial cellulose supplies the NFMs with ultrathin fiber structure, and large amounts of adsorption ligands are immobilized on the framework via the crosslinking with antibacterial ions. Thus, different from other fibers, HA-PAO NFMs achieve ultrathin diameter (20-30 nm), high BET area (51 m2 g-1), and excellent mechanical strength (13.6 MPa). The uranium adsorption capacity reaches to 409 mg-U/g-Ads in the simulated seawater, 99.2% uranium can be removed from the U-contained wastewater, and the adsorption process can be observed by the naked eye due to the significant color changes. The inhibition zones indicate their excellent anti-biofouling ability, which contributes to 1.83 times more uranium extraction amount from natural seawater than the non-antifouling adsorbents. Furthermore, they display a long service life and can be large-scale prepared, and the HA-PAO NFMs have potential in the massive uranium recovery.

Spatiotemporal tracking of the transport of RNA nano-drugs: from transmembrane to intracellular delivery.

The popularity of RNA nanoparticles (RNPs) has risen rapidly during the past decade due to the development of RNA nanotechnology. Understanding the fast dynamic process of cell entry and intracellular delivery of RNPs is essential for the design of intelligent therapeutic RNA nano-drugs and mRNA vaccines.How the interaction between the membrane and target ligand of RNPs influences the cell entry, and how the dynamic mechanism of RNPs takes place in different organelles remain ill-defined. Herein, the cell entry of Antimir21-RNP-Apt is monitored using a force tracing technique with a high spatiotemporal resolution at the single particle level, the specific interaction of Apt and EGFR promotes the cell entry efficiency and achieves long-lasting curative effects. Furthermore, the intracellular delivery pathway through different organelles is discovered using fluorescence tracking, and the low motility in early endosomes and the high motility in late endosomes are analyzed. This report provides key strategies for engineering RNA nanomedicines and facilitating clinical translation.

Nucleation and growth of elastin-like peptide fibril multilayers: an in situ atomic force microscopy study.

Controlling how molecules assemble into complex supramolecular architectures requires careful consideration of the subtle inter- and intra-molecular interactions that control their association. This is particularly crucial in the context of assembly at interfaces, where both surface chemistry and structure can play a role in directing structure formation. We report here the results of a study into the self-assembly of the elastin-like peptide EP I on structurally modified highly ordered pyrolytic graphite, including the role of spatial confinement on fibril nucleation and the growth of oriented fibril multilayers. In situ atomic force microscopy performed in fluid and at elevated temperature provided direct evidence of frustrated fibril nuclei and oriented growth of independent fibril domains. These results portend the application of this in situ strategy for studies of the nucleation and growth mechanisms of other fibril- and amyloid-forming proteins.

Neglected parasite reservoirs in wetlands: Prevalence and diversity of avian haemosporidians in waterbird communities in Northeast China.

The diversity of waterbirds is threatened, and haemosporidian parasite infection is considered one of the most important causative factors. However, to date, only a few studies focusing on specific parasite species have been carried out, which cannot reflect the general patterns at the community level. To test whether the reported haemosporidian diversity in waterbirds is underestimated, we estimated the prevalence and lineage diversity of avian haemosporidian parasites in 353 waterbirds from 26 species in the Tumuji National Nature Reserve, Northeast China, as well as the host-parasite associations. According to the molecular analysis of cytochrome b (cyt b) barcode sequences, 28.3% of the birds were infected by 49 distinct parasite lineages, including 11 Plasmodium, 12 Haemoproteus, and 26 Leucocytozoon lineages, of which 39 were novel. The highest prevalence was contributed by Leucocytozoon (13.31%), followed by Plasmodium (13.03%) and Haemoproteus (4.25%), which suggested that waterbirds were infected to a lesser extent by Haemoproteus than by the other two genera. Among the most sampled birds, species belonging to Anatidae appeared to be susceptible to Leucocytozoon but resistant to Plasmodium, while Rallidae presented the opposite pattern. On the phylogenetic tree, most of the Leucocytozoon lineages detected in Anatidae clustered together and formed two well-supported clades, while lineages restricted to Gruidae were distantly related to other parasites in all three genera. SW5 was the most abundant lineage and therefore might be a major threat to waterbirds; among the hosts, the common coot harboured the highest diversity of parasite lineages and thus could act as a reservoir for potential transmission. This is the first study of avian haemosporidian infections in a wild waterbird community in Asia. Our findings have doubled the number of lineages recorded in waterbirds, broadened our understanding of host-parasite associations, and addressed the importance of studying haemosporidian infections in wild waterbird conservation.