Energetic Lifestyle Drives Size and Shape of Avian Erythrocytes.

The size and shape of red blood cells (erythrocytes) is determined by key life history strategies in vertebrates. They have a fundamental role to deliver oxygen to tissues, and their ability to do so is shaped by the tissue's need and their shape. Despite considerable interest in how other components of blood are shaped by ecology and life history, few studies have considered erythrocytes themselves. We tested how erythrocyte size and shape varied in relation to energetically demanding activities using a dataset of 631 bird species. We found that in general, birds undergoing greater activities such as long distance migration had smaller and more elongated cells, while those with greater male-male competition had smaller and rounder cells. Smaller, more elongated erythrocytes allow more rapid oxygenation/deoxygenation and support greater aerobic activity. The rounder erythrocytes found in species with strong male-male competition may stem from younger erythrocytes deriving from androgen-induced erythropoiesis rates. Finally, diving species of bird had larger erythrocytes, indicating that erythrocytes are acting as a vital oxygen store. In summary, erythrocyte size and shape in birds are driven by the need to deliver oxygen during energetically costly activities.

Bactericidal activity of avian complement: a contribution to understand avian-host tropism of Lyme borreliae.

Complement has been considered as an important factor impacting the host-pathogen association of spirochetes belonging to the Borrelia burgdorferi sensu lato complex, and may play a role in the spirochete's ecology. Birds are known to be important hosts for ticks and in the maintenance of borreliae. Recent field surveys and laboratory transmission studies indicated that certain avian species act as reservoir hosts for different Borrelia species. Nevertheless, our current understanding of the molecular mechanisms determining host tropism of Borrelia is still in its fledgling stage. Concerning the role of complement in avian-host tropism, only a few bird species and Borrelia species have been analysed so far. Here, we performed in vitro serum bactericidal assays with serum samples collected from four bird species including the European robin Erithacus rubecula, the great tit Parus major, the Eurasian blackbird Turdus merula, and the racing pigeon Columba livia, as well as four Borrelia species (B. afzelii, B. garinii, B. valaisiana, and B. burgdorferi sensu stricto). From July to September 2019, juvenile wild birds were caught using mist nets in Portugal. Racing pigeons were sampled in a loft in October 2019. Independent of the bird species analysed, all Borrelia species displayed an intermediate serum-resistant or serum-resistant phenotype except for B. afzelii challenged with serum from blackbirds. This genospecies was efficiently killed by avian complement, suggesting that blackbirds served as dead-end hosts for B. afzelii. In summary, these findings suggest that complement contributes in the avian-spirochete-tick infection cycle and in Borrelia-host tropism.

Revisiting the question of nucleated versus enucleated erythrocytes in birds and mammals.

Erythrocyte enucleation is thought to have evolved in mammals to support their energetic cost of high metabolic activities. However, birds face similar selection pressure yet possess nucleated erythrocytes. Current hypotheses on the mammalian erythrocyte enucleation claim that the absence of cell organelles allows erythrocytes to 1) pack more hemoglobin into the cells to increase oxygen carrying capacity and 2) decrease erythrocyte size for increased surface area-to-volume ratio, and improved ability to traverse small capillaries. In this article, we first empirically tested current hypotheses using both conventional and phylogenetically informed analysis comparing literature values of mean cell hemoglobin concentration (MCHC) and mean cell volume (MCV) between 181 avian and 194 mammalian species. We found no difference in MCHC levels between birds and mammals using both conventional and phylogenetically corrected analysis. MCV was higher in birds than mammals according to conventional analysis, but the difference was lost when we controlled for phylogeny. These results suggested that avian and mammalian erythrocytes may employ different strategies to solve a common problem. To further investigate existing hypotheses or develop new hypothesis, we need to understand the functions of various organelles in avian erythrocytes. Consequently, we covered potential physiological functions of various cell organelles in avian erythrocytes based on current knowledge, while making explicit comparisons with their mammalian counterparts. Finally, we proposed by taking an integrative and comparative approach, using tools from molecular biology to evolutionary biology, would allow us to better understand the fundamental physiological functions of various components of avian and mammalian erythrocytes.

HEALTH STATUS OF NAZCA BOOBIES (SULA GRANTI) ON DAPHNE MAJOR ISLAND IN THE GALÁPAGOS DETERMINED BY HEMATOLOGY, BIOCHEMISTRY, AND PHYSICAL EXAMINATION.

Island species are particularly vulnerable to environmental disturbances and introduced pathogens. Conducting health assessments of wild populations in the Galápagos improves the ability of wildlife managers and veterinarians to detect deteriorations in health status. Seabirds in particular are useful species to monitor due to their colonial breeding and wide migration range. Nazca boobies (Sula granti) in a breeding colony at Daphne Major (n = 30) were given physical examinations, and blood samples were collected for hematology and biochemistry using an iSTAT Portable Clinical Analyzer. Female boobies had longer wing length than males, as well as lower blood urea nitrogen, creatinine, and white blood cell counts. This could be attributed to sexual dimorphism or differences in foraging and mating strategies between the sexes. The time between capture and blood collection had a significant inverse relationship on plasma sodium, potassium, hemoglobin, anion gaps, and lymphocyte counts, suggesting that blood sampling in Nazca boobies should be done in less than 5 m to avoid the impacts of stress on hematological parameters. This is the first health assessment on the breeding colony of Nazca boobies at Daphne Major, and the results can inform future monitoring in this species as well as other sulids.

Description of Haemoproteus asymmetricus n. sp. (Haemoproteidae), with remarks on predictability of the DNA haplotype networks in haemosporidian parasite taxonomy research.

Haemoproteus species (Haemosporida, Haemoproteidae) are cosmopolitan blood parasites, which have been neglected for over 100-years, but attracted attention recently due to reports of severe and even lethal haemoproteosis in birds and vectors. Approximately 150 species of avian Haemoproteus have been described and named, but molecular data suggest that hundreds of independently evolving molecular lineages might occur, indicating the existence of a remarkable undescribed species diversity. It is timely to develop a methodology, which allow the application of available genetic data in taxonomy of haemosporidians on species levels. This study aimed to test a hypothesis suggesting that DNA haplotype networks might aid in targeting genetically distinct, but still undescribed parasites, and might be used to direct taxonomic studies on haemosporidian species levels. Mainly, we tested a prediction that the lineage hTUPHI01, a common Haemoproteus parasite of Turdus philomelos, might be a new species, which is morphologically similar and genetically closely related to the parasites of Haemoproteus minutus group. Blood samples of T. philomelos naturally infected with this parasite lineage were collected and studied using microscopic examination of blood films and PCR-based methods. Haemoproteus asymmetricus n. sp. was found in this bird, described and characterised molecularly using partial cytochrome b (cytb) sequences. The new species shared some features with parasites of the H. minutus group, as was predicted by the DNA haplotype network. Due to the visualisation of closely related lineages as well as the evaluation of their host and geographic distributions, DNA haplotype networks can be recommended as the helpful methodology, able to direct and speed practical work on parasite species taxonomy and pathogen biodiversity. The combined molecular phylogenetic and morphological approaches showed that the well-supported clades in Bayesian phylogenetic trees based on the partial cytb gene sequences contain morphologically remarkably different Haemoproteus parasite species, which however, share some basic biological features. Phylogenetic analysis can be used for prediction of these basic features in still undescribed parasites. This study calls for further fusion of advanced molecular and microscopy approaches for better understanding haemosporidian parasite biology.

Erythroid enucleation: a gateway into a "bloody" world.

Erythropoiesis is an intricate process starting in hematopoietic stem cells and leading to the daily production of 200 billion red blood cells (RBCs). Enucleation is a greatly complex and rate-limiting step during terminal maturation of mammalian RBC production involving expulsion of the nucleus from the orthochromatic erythroblasts, resulting in the formation of reticulocytes. The dynamic enucleation process involves many factors ranging from cytoskeletal proteins to transcription factors to microRNAs. Lack of optimum terminal erythroid maturation and enucleation has been an impediment to optimum RBC production ex vivo. Major efforts in the past two decades have exposed some of the mechanisms that govern the enucleation process. This review focuses in detail on mechanisms implicated in enucleation and discusses the future perspectives of this fascinating process.

Seasonal variation in sex-specific immunity in wild birds.

Whilst the immune system often varies seasonally and exhibits differences between males and females, the general patterns in seasonality and sex differences across taxa have remained controversial. Birds are excellent model organisms to assess these patterns, because the immune system of many species is well characterised. We conducted a meta-analysis using 41 wild bird species from 24 avian families to investigate sex differences and seasonal (breeding/non-breeding) variations in immune status, including white blood cell counts, phytohaemagglutinin (PHA) test, bacteria-killing ability (BKA), haemolysis and haemagglutination assays. We found male-biased macrophage concentration, BKA and haemolysis titers, but only during the breeding season. Sex-specific heterophil concentrations, heterophil/lymphocyte ratios and PHA responses differed between breeding and non-breeding, suggesting larger changes in males than in females. Importantly, sex differences in immune status are stronger during the breeding period than during the non-breeding period. Taken together, our study suggests that both seasonal variation and sex differences in immune system are common in birds, although their associations are more complex than previously thought.

Blood parasites of bird communities in Sri Lanka and their mosquito vectors.

Avian blood parasites have been shown to have significant health effects on avifauna worldwide. Sri Lanka, a tropical island rich with resident and migratory birds, has not been properly evaluated for avian blood parasites or their vectors. We investigated the presence of avian haemoparasites in Sri Lankan birds and the potential mosquito vectors of those pathogens. Blood samples were collected from local/migratory birds captured by standard mist nets from Anawilundawa bird sanctuary, Hanthana mountain range, and the University of Peradeniya park. Mosquitoes were collected from Halgolla forest reserve and the forest patches in Kurunegala and Gampola areas in addition to the above mist-netting localities. Part of the mitochondrial cytochrome b (cytb) gene was amplified and sequenced to detect the presence of haemoparasites from avian blood samples (86) and mosquito samples (480). Blood parasites of the two genera, i.e., Haemoproteus (4 species; Haemoproteus sp. 1-4) and Plasmodium (5 species; Plasmodium sp. 1-5) were identified from seven bird species (four resident and three migratory). Among these, three bird species (Red-vented bulbul (3/16), Asian Brown flycatcher (1/1), and India pitta (1/1)) were positive for Plasmodium spp., while four (Yellow-browed bulbul (1/4), oriental white-eye (1/4), brown-headed Barbet (1/4), and Indian blue robin (1/1)) were positive for Haemoproteus spp. Two mosquito species were also positive for Plasmodium (3) and Haemoproteus (1) species. Phylogenetic analysis and haplotype networks created using positive sequences of haemoparasites showed that a Plasmodium clade was shared by Cx nigropunctatus mosquitoes and the migratory bird, Indian pitta. The majority (85%) of the Plasmodium and Haemoproteus sequences of this study were not linked to the well-characterized species suggesting the distinct nature of the lineages. Associations between mosquito species and blood parasites of birds suggest the possible vector status of these mosquitoes.

CLINICAL CHEMISTRY, VITAMIN, ELECTROPHORESIS, AND HEMATOLOGIC ANALYTES OF BLACK-HEADED IBIS (THRESKIORNIS MELANOCEPHALUS).

The evaluation of laboratory blood analytes is an important tool for health assessment in avian medicine. Unfortunately, there is a lack of suitable reference values for many zoo bird species. The goal of the present study was to establish reference intervals for a wide range of blood analytes in healthy black-headed ibis (Threskiornis melanocephalus) kept in a zoologic garden. Lithium heparinized blood samples from 15 individuals were collected, and 18 different clinical chemistry analytes were measured. New are especially the minimum and maximum values for bile acid (2.38-49.4 µmol/L), cholinesterase (439-1260 U/L), vitamin A (0.65-1.70 mg/L), vitamin E (26.3-52.5 mg/L), and capillary zone electrophoresis (prealbumin, 10.2-23.4%; albumin, 35.9-44.1%; A/G ratio, 1.07-1.69; α-, 15.7-20.0%; ß-, 13.5-19.1%; γ-globulin, 6.08-11.3%). The measured values for clinical chemistry, vitamin, electrophoresis, and hematologic analytes are a basis for further studies and for diagnostics and clinical treatment in this ibis species.

Prognostic Indicators of Avian Survival.

The objective of this study was to evaluate whether blood glucose, packed cell volume, and total protein are effective diagnostic tests to predict short-term (48 h following admittance to a veterinary hospital) prognosis for wild birds with traumatic injuries. The cohort study conducted for this investigation took place at the Jivdaya Charitable Trust in Ahmedabad, India, over a 7-d period, at the time of the International Kite Flying Festival of Uttarayan. A small blood sample was collected from every avian patient prior to surgical treatment and immediately analyzed. Patient outcome was reviewed and recorded at the end of a 48-h period following admittance to the veterinary hospital. Statistical analysis revealed no significant correlations between blood glucose levels (P = .24), packed cell volume (P = .80), or total protein levels (P = .47) outside the reference intervals and short-term outcome of the patients. There is a lack of evidence regarding the use of these diagnostic tests to ascertain short-term prognosis in avian species. While research has been conducted on avian wound healing, studies fail to identify any correlation between duration of injury and patient outcome. Despite a lack of statistically significant results from this study, the findings should not be dismissed and could be used as a basis for future studies on this subject.

Molecular Analysis of the Bloodmeals of Culex spp. Mosquitoes at Natural Habitats in Singapore to Investigate the Potential Risk of Japanese Encephalitis Virus and West Nile Virus Transmission.

Japanese encephalitis virus (JEV) and West Nile virus (WNV) are arboviruses primarily transmitted by Culex spp. mosquitoes. Birds are the primary hosts for JEV and WNV. Recent WNV outbreaks in Europe and United States and their association with migratory birds highlight the importance of understanding the feeding host preference of potential vectors for outbreak preparedness, especially in nonendemic settings. Singapore is nonendemic to JEV and WNV, but is a stopover site for migratory birds of the East Asian-Australasian Flyway. Therefore, we elucidated the feeding host range of Culex spp. mosquitoes captured in four natural (bird) habitats in Singapore from January 2011 to December 2012. We characterized feeding host DNA in field-caught mosquitoes using a PCR sequencing-based assay targeting the mitochondrial gene regions. Of 22,648 mosquitoes captured, 21,287 belonged to the Culex vishnui subgroup. The host DNA analysis showed that mosquitoes from the Cx. vishnui subgroup are opportunistic biters, feeding on a range of birds and mammals. Cx. vishnui subgroup, Culex sitiens and Culex bitaeniorhynchus, was primarily ornithophagic, although they fed opportunistically on mammals, including humans. Culex gelidus and Culex quinquefasciatus, in contrast, fed mainly on mammals. The presence of ornitho- and anthropophilic mosquito vectors and susceptible avian and mammalian hosts poses a risk spill-over transmission of JEV and WNV among humans, should these viruses be introduced through migratory birds and establish persistent transmission in resident birds and animal hosts in Singapore.

Identification of blood-feeding sources in Panstrongylus, Psammolestes, Rhodnius and Triatoma using amplicon-based next-generation sequencing.

BACKGROUND: Triatomines are hematophagous insects that play an important role as vectors of Trypanosoma cruzi, the causative agent of Chagas disease. These insects have adapted to multiple blood-feeding sources that can affect relevant aspects of their life-cycle and interactions, thereby influencing parasitic transmission dynamics. We conducted a characterization of the feeding sources of individuals from the primary circulating triatomine genera in Colombia using amplicon-based next-generation sequencing (NGS). METHODS: We used 42 triatomines collected in different departments of Colombia. DNA was extracted from the gut. The presence of T. cruzi was identified using real-time PCR, and discrete typing units (DTUs) were determined by conventional PCR. For blood-feeding source identification, PCR products of the vertebrate 12S rRNA gene were obtained and sequenced by next-generation sequencing (NGS). Blood-meal sources were inferred using blastn against a curated reference dataset containing the 12S rRNA sequences belonging to vertebrates with a distribution in South America that represent a potential feeding source for triatomine bugs. Mean and median comparison tests were performed to evaluate differences in triatomine blood-feeding sources, infection state, and geographical regions. Lastly, the inverse Simpson's diversity index was calculated. RESULTS: The overall frequency of T. cruzi infection was 83.3%. TcI was found as the most predominant DTU (65.7%). A total of 67 feeding sources were detected from the analyses of approximately 7 million reads. The predominant feeding source found was Homo sapiens (76.8%), followed by birds (10.5%), artiodactyls (4.4%), and non-human primates (3.9%). There were differences among numerous feeding sources of triatomines of different species. The diversity of feeding sources also differed depending on the presence of T. cruzi. CONCLUSIONS: To the best of our knowledge, this is the first study to employ amplicon-based NGS of the 12S rRNA gene to depict blood-feeding sources of multiple triatomine species collected in different regions of Colombia. Our findings report a striking read diversity that has not been reported previously. This is a powerful approach to unravel transmission dynamics at microgeographical levels.

Female birdsong.

Naomi Langmore introduces female song in birds.

Validation of Multi-Residue Method for Quantification of Antibiotics and NSAIDs in Avian Scavengers by Using Small Amounts of Plasma in HPLC-MS-TOF.

Pharmaceuticals are still considered emerging pollutants affecting both aquatic and terrestrial ecosystems. Scavenging bird species may be exposed to veterinary drugs when they feed on livestock carcasses provided at supplementary feeding stations, as these are often stocked with ailing and/or recently medicated animals. Because those animals may be a source of several different pharmaceutical compounds, analytical methods to evaluate residue levels and exposure potential should enable detection and quantification of as many different compounds as possible, preferably from small sample volumes. Four different extraction methods were tested to conduct HPLC-MS-TOF analysis of some of the most common veterinary drugs used in livestock in Spain. The method deemed most viable was a simple extraction, using methanol and 100 µL of plasma, that allowed quantification of seven antibiotics (tetracycline, oxytetracycline, ciprofloxacin, enrofloxacin, nalidixic acid, trimethoprim, sulfadiazine) and five nonsteroidal anti-inflammatory drugs (NSAIDs) (meloxicam, flunixin, carprofen, tolfenamic acid, phenylbutazone). The method was then applied to analysis of 29 Eurasian griffon vulture (Gyps fulvus) nestling samples, wherein enrofloxacin and tolfenamic acid were most commonly detected (69% and 20%, respectively). To our knowledge, this is the first study including NSAIDs in the exposure assessment of different classes of veterinary pharmaceuticals in live avian scavengers.

Anti-microbial activity of whole blood and plasma collected from Anna's Hummingbirds (Calypte anna) against three different microbes.

Hummingbirds are essential pollinators in many ecosystems, making their conservation critical. As is the case with many species, hummingbirds are now facing a variety of challenges resulting from anthropogenic changes. As populations shift and species interactions change, disease is likely to pose a significant threat. There is a basic understanding of which pathogens currently affect a variety of hummingbird species, however there is a paucity of information about their immune systems capacity to kill pathogens and what specific factors may affect immunity. The objective of this study was to gain a basic understanding of the effect of age, sex, and molt on the constitutive innate immunity of hummingbirds. An in vitro assay was used to assess the microbiocidal capacity of the whole blood of Anna's Hummingbirds (Calypte anna) against three different microbes: Escherichia coli (E. coli), Staphylococcus aureus (S. aureus) and Candida albicans (C. albicans). The effect of age, sex and molt on anti-microbial capacity varied based on the microbe type. After-hatch-year birds tended to have better anti-microbial capacity compared to hatch-year birds. Male birds had higher anti-microbial activity than female birds, although this was not observed against C. albicans. Molting birds had a weaker antimicrobial activity against E. coli and S. aureus than birds that were not molting. These results represent an important first step towards defining the parameters of constitutive innate immunity of Anna's Hummingbirds as well as providing important knowledge about factors that should be considered when evaluating the health of wild populations.

Organ Histopathology and Hematological Changes Associated With Heat Exposure in Australian Desert Birds.

An inability of the body to appropriately respond to extreme temperatures will result in pathological changes to vital organs and adverse hematological changes. Mild heat exposure of a bird to a temperature above the zone of thermoneutrality can induce subclinical heat stress, which may be a precursor to illness. The ability to identify subtle changes that may be associated with subclinical heat stress can be important in early diagnosis and treatment of heat stress in birds. Pathological changes to internal body organs, post-heat exposure, were microscopically examined in 13 budgerigars (Melopsittacus undulatus), 15 zebra finches (Taeniopygia guttata), and 8 diamond doves (Geopelia cuneata) as model species for the bird orders Psittaciformes, Passeriformes, and Columbiformes, respectively. There was mild to moderate congestion of the lungs of 28/36 birds examined, including all of the budgerigars and diamond doves. In 8/15 zebra finches no significant lung congestion was noted. Interstitial and pulmonary hemorrhage was in observed in one diamond dove. The most common hepatic pathologic change identified was micro- and macro-vesicular hepatocellular vacuolation (4/15 zebra finches, 5/13 budgerigars, and 8/8 diamond doves). There was mild to moderate congestion in the kidneys of 1/15 zebra finch, 2/ 13 budgerigars, and 4/8 diamond doves, as well as in the gastrointestinal tract of 1/15 zebra finch and 7/8 budgerigars. Budgerigars showed a decrease in hematocrit and a significant change in the numbers of heterophils and lymphocytes following heat exposure. The basophil population of cells remained relatively stable in both budgerigars and diamond doves. These findings indicate avian species differences in body organ and hematological changes following exposure to similar elevations in environmental temperatures.

Wild Bird Surveillance for Avian Influenza Virus.

Avian influenza (AI) viruses have been routinely isolated from a wide diversity of free-living avian species, representing numerous taxonomic orders. Birds in orders Anseriformes and Charadriiformes are considered the natural reservoirs for all AI viruses; it is from these orders that AI viruses have been most frequently isolated. Since first recognized in the late 1800s, AI viruses have been an important cause of disease in poultry and, occasionally, in non-gallinaceous birds and mammals. While AI viruses tend to be of low pathogenicity (LP) in wild birds, the 2014-2015 incursion of highly pathogenic avian influenza (HPAI) clade 2.3.4.4 H5Nx viruses into North America and the recent circulation of HPAI H5 viruses in European wild birds highlight the need for targeted, thorough, and continuous surveillance programs in the wild bird reservoir. Such programs are crucial to understanding the potential risk for the incursion of AI into human and domestic animal populations. The aim of this chapter is to provide general concepts and guidelines for the planning and implementation of surveillance plans for AI viruses in wild birds.

Avian Influenza Virus Sample Types, Collection, and Handling.

Successful detection of avian influenza (AI) virus, viral antigen, nucleic acid, or antibody is dependent upon the collection of the appropriate sample type, the quality of the sample, and the proper storage and handling of the sample. The diagnostic tests to be performed should be considered prior to sample collection. Sera are acceptable samples for ELISA or agar gel immunodiffusion tests, but not for real-time RT-PCR. Likewise, swabs and/or tissues are acceptable for real-time RT-PCR and virus isolation. The sample type will also depend on the type of birds that are being tested; oropharyngeal swabs from gallinaceous poultry and cloacal swabs from waterfowl are the preferred specimens for most diagnostic tests, although it is optimal to collect swabs from both locations, if possible. In addition to collecting the appropriate sample for the tests to be performed, selecting the right materials for sample collection (i.e., type of swab) is very important. This chapter will outline the collection of different specimen types and procedures for proper specimen handling.

The buffy coat method: a tool for detection of blood parasites without staining procedures.

BACKGROUND: Blood parasites belonging to the Apicomplexa, Trypanosomatidae and Filarioidea are widespread in birds and have been studied extensively. Microscopical examination (ME) of stained blood films remains the gold standard method for the detection of these infections in birds, particularly because co-infections predominate in wildlife. None of the available molecular tools can detect all co-infections at the same time, but ME provides opportunities for this to be achieved. However, fixation, drying and staining of blood films as well as their ME are relatively time-consuming. This limits the detection of infected hosts during fieldwork when captured animals should be released soon after sampling. It is an obstacle for quick selection of donor hosts for parasite experimental, histological and other investigations in the field. This study modified, tested and described the buffy coat method (BCM) for quick diagnostics (~ 20 min/sample) of avian blood parasites. METHODS: Blood of 345 birds belonging to 42 species was collected, and each sample was examined using ME of stained blood films and the buffy coat, which was examined after centrifugation in capillary tubes and after being transferred to objective glass slides. Parasite detection using these methods was compared using sensitivity, specificity, positive and negative predictive values and Cohen's kappa index. RESULTS: Haemoproteus, Leucocytozoon, Plasmodium, microfilariae, Trypanosoma and Lankesterella parasites were detected. BCM had a high sensitivity (> 90%) and specificity (> 90%) for detection of Haemoproteus and microfilariae infections. It was of moderate sensitivity (57%) and high specificity (> 90%) for Lankesterella infections, but of low sensitivity (20%) and high specificity (> 90%) for Leucocytozoon infections. Trypanosoma and Plasmodium parasites were detected only by BCM and ME, respectively. According to Cohen's kappa index, the agreement between two diagnostic tools was substantial for Haemoproteus (0.80), moderate for Lankesterella (0.46) and fair for microfilariae and Leucocytozoon (0.28) infections. CONCLUSIONS: BCM is sensitive and recommended as a quick and reliable tool to detect Haemoproteus, Trypanosoma and microfilariae parasites during fieldwork. However, it is not suitable for detection of species of Leucocytozoon and Plasmodium. BCM is a useful tool for diagnostics of blood parasite co-infections. Its application might be extended to studies of blood parasites in other vertebrates during field studies.

Veterinary utility of dried blood spots for detailed analysis of chlorinated pesticides and polychlorinated biphenyls by gas chromatography tandem mass spectrometry.

Persistent organic pollutants (POPs) are organic compounds of anthropogenic origin that resist atmospheric and microbial degradation and thus persist in the environment and in food chains for exceptionally long periods of time. Veterinarians and wildlife researchers need simple methodologies for monitoring and measuring such compounds including two large and diverse categories, organochlorine pesticides (OCs) and polychlorinated biphenyls (PCBs), compounds that have been largely banned from production and use except for specific exceptions. We present development of methodologies for detection and quantitation of 22 OCs and 10 PCB congeners by tandem quadrupole gas chromatography-mass spectrometric analysis of Dried Blood Spots (DBS). Development was enabled by (1) optimization of suspension and extraction methodologies for DBS; (2) strategic streamlining and condensation of Multiple Reaction Monitoring (MRM) settings on GC/MS/MS; and (3) improvement of GC settings to accommodate all 32 compounds in a single chromatographic run per sample. The method was validated for parameters of linearity, limits of detection and quantitation, recovery and precision, and results from blood were shown to correlate well with those from DBS despite both being only 50 uL in volume. The method was applied successfully to blood samples from nine avian specimens submitted to the MSU Veterinary Diagnostic Lab, and all were shown to bear the burden of varying levels of OCs and/or PCB compounds.