Enantioselective total synthesis and biological evaluation of (-)-solanacol.

An enantioselective synthesis of the phenyl ring-containing strioglactone, (-)-solanocol, is described. Application of a Dynamic Kinetic Resolution (DKR) in the stereo-defining step enabled a step-economical synthesis to be achieved, and allowed access to natural and non-natural enantiomers with equal facility. Results of seed germination assays and Differential Scanning Fluorimetry (DSF) measurements with the known strigolactone receptor protein, Decreased Apical Dominance 2 (DAD2), are reported.

Heterobilharzia americana infection and congestive heart failure in a llama (Lama glama).

The schistosome Heterobilharzia americana infects several mammalian species in the southeastern United States, including horses, but infections have not been reported in camelids. This is a report of H. americana infection in a 6-year-old llama with extensive cardiac pathology and congestive heart failure. Parasite-induced granulomas were widely disseminated and included overwhelming involvement of the lungs and liver. Microscopic lesions in the heart included myofiber degeneration and necrosis, with extensive replacement fibrosis. Polymerase chain reaction amplification and sequencing confirmed the presence of H. americana in the lungs.

No change in hemoglobin mass after 40 days of physical activity in previously untrained adults.

A high hemoglobin mass (Hb(mass)) is associated with a high maximum aerobic power (VO(2max)), however, the extent to which Hb(mass) is influenced by training is currently unclear. Accordingly, this study monitored changes in Hb(mass) and VO(2max) in 12 previously untrained adults (aged 18-25 years) following 40 days of regular physical activity. Hb(mass) and VO(2max) were assessed at the start and end of a 40-day physical activity program, which comprised of approximately 40 min of daily, moderate-intensity physical activity. Relative VO(2max) increased by 11.3%, yet there was no significant change in relative Hb(mass) (1.7%) and body mass (0.2%) during the 40-day period. There was a significant correlation between Hb(mass) and VO(2max) at the start of the study (r=0.58, P=0.05), but not between the change in relative VO(2max) and the change in relative Hb(mass) (r=-0.07, P=0.83). Our results support the concept of relative stability in Hb(mass) with approximately 1 month of moderate-intensity physical activity suggesting that Hb(mass) may be used for talent identification and possibly for anti-doping purposes.

Natural Heterobilharzia americana infection in horses in Texas.

The schistosome Heterobilharzia americana infects dogs, raccoons, and other mammals in the southeastern United States. Migration of eggs into the liver results in parasitic granulomas with varying degrees of fibrosis and inflammation. Recently, hepatic parasitic granulomas in horses were shown to be caused by H. americana infection. In the present study, samples of liver from 11 of 12 horses with hepatic granulomas identified at necropsy (n = 11) or surgical biopsy (n = 1) were used for DNA extraction, polymerase chain reaction amplification and sequencing using primers specific for a portion of the H. americana small subunit ribosomal RNA gene. A polymerase chain reaction amplicon of the correct size was produced from the extracted DNA in 8 of the 11 horses. Amplicons from 5 of the 8 positive horses were sequenced and had 100% identity with H. americana. In all but 2 of the 12 horses, Heterobilharzia was not responsible for the primary clinical disease, and the hepatic granulomas were considered an incidental finding.

Detraining decreases Hb(mass) of triathletes.

Haemoglobin mass (Hbmass) determination using CO rebreathing may assist to detect illegal blood doping practices, however variations in Hbmass with periods of intensive training and detraining must be quantified. This study aimed to determine the effect of a 30-day period of detraining on Hbmass in ultra-endurance triathletes. 9 male recreational triathletes (29-44 years) participated in the study. Hbmass was assessed using CO rebreathing 30 days and 10 days before an ultra-endurance triathlon and after ~10, 20 and 30 days of detraining following the race. V˙O2max was assessed 10 days before the race and also after the 30-day detraining period, which consisted of an 87% reduction in training hours. After 30-days of detraining there was a 3.1% decrease in mean Hbmass from 868±99 to 840±94 g, (p=0.03), and a 4.7% decrease in mean V˙O2max from 4.83±0.29 to 4.61±0.41 L/min as well as a 2.8% increase of body mass from 75.1±6.4 to 77.1±6.1 kg and a 28% increase in skinfold total from 43.9±14.2 to 55.1±14.0 mm. Individual decreases in Hbmass following detraining would need to be considered if using Hbmass for anti-doping purposes.

Heterobilharzia americana infection in a Grant's zebra (Equus burchelli boehmi).

Heterobilharzia americana is a trematode parasite (family: Schistosomatidae) of mammalian wildlife that occasionally infects domestic dogs and horses in the southeastern United States. This report presents the first case of H. americana infection in a Grant's zebra (Equus burchelli boehmi). The parasite was diagnosed post mortem as an incidental finding by histopathologic identification of H. americana eggs in the liver parenchyma after the zebra succumbed in an accidental traumatic death. Molecular analysis of a portion of the parasite small subunit ribosomal RNA gene demonstrated 100% identity with DNA sequences from parasites previously characterized in domestic horses. Equine infections with H. americana are considered uncommon, but are probably underdiagnosed.

Eight cases of feline cutaneous leishmaniasis in Texas.

Leishmaniasis is a zoonotic disease caused by intracellular Leishmania protozoa that are transmitted by sandflies. The disease occurs in 3 forms: cutaneous, mucocutaneous, and visceral. Cutaneous leishmaniasis has been reported in cats in Europe and South America and in 1 cat from Texas. Leishmania mexicana is endemic in Texas and has been reported to cause cutaneous lesions in humans. This article describes the pathology of 8 biopsy cases of feline cutaneous leishmaniasis presented to the Texas Veterinary Medical Diagnostic Laboratory over a 3.5-year period. The median age of the cats was 3 years; each was presented with nodular, ulcerative lesions on the pinnae and less commonly on the muzzle and periorbital skin. Histologically, the lesions were nodular to diffuse histiocytic dermatitis with numerous amastigotes (2-4 µm) within macrophages and occasionally within the interstitium. Organisms were often contained within round, clear, intracellular vacuoles. In areas of necrosis, organisms were also free within the interstitium. The overlying epidermis was hyperkeratotic, hyperplastic, and often ulcerated. The organisms were not argyrophilic (Gomori methenamine silver), reacted poorly with periodic acid-Schiff reagent, and were inconsistently basophilic with Giemsa. Although not readily visible histologically, kinetoplasts were evident in amastigotes in cytologic preparations. The lesions were similar to those described for cutaneous L. mexicana infection in humans. In 5 of the 8 cats, Leishmania mexicana DNA was amplified from paraffin-embedded tissue by polymerase chain reaction and sequenced.

Distribution and characterization of canine Chagas disease in Texas.

Although acute and chronic cases of canine Chagas disease have been reported from multiple areas in the southern region of the United States, little data are available on current disease occurrence patterns in endemic areas. Therefore, a study to assess frequency, geographic distribution, signalment, and clinical spectrum of Chagas disease in domestic dogs from Texas was conducted. Serology, histopathology, and clinical case records from multiple institutions for the time period 1993-2007 were analyzed. A total of 537 serologically and/or histopathologically confirmed cases were documented. Cases were reported from 48 of 254 counties within Texas, covering all major geographic regions. Forty-eight dog breeds were represented among the cases, primarily in the sporting and working groups. In histopathologically confirmed cases, acute death occurred in 42%, approximately half of which were <1 year of age. Nearly all cases with histopathology data reported myocarditis (97.9%) and observation of Trypanosoma cruzi organisms (81.7%). Predominant clinical observations included enlarged heart, lethargy, anorexia, ascites, cardiac conduction disturbances, and respiratory difficulties. An increasing rate of serologic test submissions was noted over the study period, with an overall positive test prevalence of 20.3%. The study results provide strong evidence that an active canine Chagas disease transmission cycle is present throughout all ecoregions of Texas, affecting a broad range of dog breeds and age groups.

Trypanosoma cruzi discrete typing unit TcIV implicated in a case of acute disseminated canine Chagas disease.

In 2006, Nabity et al. reported on an atypical presentation of Trypanosoma cruzi (T. cruzi) infection in an 8-month old English Mastiff from central Texas. Clinical signs and laboratory findings included lymphadenopathy, weight loss, amastigotes in lymph node aspirates, and initial serological results suggestive of either T. cruzi or Leishmania infection. Given the poor prognosis, the dog was euthanized and subsequent testing and culture of parasites from a lymph node revealed T. cruzi infection. Because different parasite discrete typing units (DTUs) are potentially associated with different disease outcomes in a variety of mammalian hosts, an understanding of these relationships in naturally infected dogs may be useful for informing canine prognosis and may also have human health implications. Here, we compared strains using culture versus culture-independent methods. We subjected archived cultured parasites harvested from the lymph node in the infected Mastiff to two independent approaches for determining parasite DTU, including sequencing of the TcSC5D gene and use of DTU-specific qPCR probes to hybridize the nuclear spliced leader intergenic region (SL-IR). Both approaches revealed T. cruzi discrete typing unit TcIV. Testing of multiple other tissues directly without culturing, including heart/tongue, intestine, trachea/lymph nodes, and uterus/ovary, provided further evidence of disseminated TcIV infection in this dog. We report T. cruzi DTU TcIV as the cause of a severe disseminated infection in a dog from an area with triatomine vectors in central Texas, adding to the limited body of clinicopathologic data that links specific parasite strains to disease outcomes in dogs in the US. Future studies to type parasites from asymptomatic dogs and those with diverse disease manifestations will be useful in informing the degree to which parasite genetics is associated with disease presentation and severity. If applied to antemortem samples, diagnostic typing of parasites from infected dogs may assist in determining prognosis.

Fatal encephalitozoonosis in two koalas.

Two young koalas from a fauna park, recently out of the pouch and approximately 6 months old, were found dead with no previous clinical signs or gross lesions. On histopathological examination, large numbers of spores consistent with a microsporidian organism were present intracellularly within the small intestinal mucosa. Electron microscopy and polymerase chain reaction studies (sequencing the 5' end of the SSU RNA gene) identified the organism as Encephalitozoon intestinalis with 100% homology with those of previously reported human isolates. This is believed to be the first report of this organism in a marsupial.

An atypical case of Trypanosoma cruzi infection in a young English Mastiff.

A case of Trypanosoma cruzi infection in a young English Mastiff from Texas is presented. Clinical signs and laboratory findings included subcutaneous edema, lymphadenopathy, weight loss, and hypoalbuminemia. Cytology of a lymph node revealed numerous amastigotes. No trypomastigotes were observed in buffy coat preparation of peripheral blood, and on histologic evaluation, most organs contained numerous interstitial pseudocysts. Initial serology was positive for both T. cruzi and Leishmania, and immunohistochemistry supported a diagnosis of Leishmania. However, additional serology supported a T. cruzi infection, and cultivation of organisms isolated from a lymph node revealed morphology consistent with T. cruzi. In addition, PCR analysis resulted in a 504 bp fragment with 99% homology to a flagellar protein of T. cruzi. Although uncommon, autochthonous cases of both T. cruzi and Leishmania have been reported in the United States. Clinical signs observed with both diseases can show many similarities, cytology may be indistinguishable, as in this case, and serological cross-reactivity is common. This case demonstrates an unusual presentation of T. cruzi and the use of multiple testing strategies to support its diagnosis.

Microsporidia - Emergent Pathogens in the Global Food Chain.

Intensification of food production has the potential to drive increased disease prevalence in food plants and animals. Microsporidia are diversely distributed, opportunistic, and density-dependent parasites infecting hosts from almost all known animal taxa. They are frequent in highly managed aquatic and terrestrial hosts, many of which are vulnerable to epizootics, and all of which are crucial for the stability of the animal-human food chain. Mass rearing and changes in global climate may exacerbate disease and more efficient transmission of parasites in stressed or immune-deficient hosts. Further, human microsporidiosis appears to be adventitious and primarily associated with an increasing community of immune-deficient individuals. Taken together, strong evidence exists for an increasing prevalence of microsporidiosis in animals and humans, and for sharing of pathogens across hosts and biomes.

Aluminum-Induced Genes (Induction by Toxic Metals, Low Calcium, and Wounding and Pattern of Expression in Root Tips).

We have investigated the response of four Al-induced genes (wali1, -3, -4, and -5) from wheat (Triticum aestivum L.) to other stresses. The relative transcript levels of wali1 (encoding a plant metallothionein-like protein), wali3 and wali5 (putative Bowman-Birk proteinase inhibitors), and wali4 (phenylalanine ammonialyase) increased in root tips of wheat after 2-d treatments with toxic levels of all other metals tested (Cd, Fe, Zn, Cu, Ga, In, and La). The expression levels of wali1, -3, -4, and -5 also increased in the root tips of plants grown in the presence of low levels of Ca (10[mu]M). The transcript levels of wali1, -3, and -5 increased in wounded leaf and root tissue, whereas the transcript levels of wali4 increased only in wounded leaves. The site of expression of wali1, -3, and -5 in root tips was identified using in situ hybridization. wali1 was expressed predominantly in the meristematic tissue of the root tip, whereas wali3 and wali5 were expressed predominantly in the cortical tissue of the root. Some changes in the site of expression of these genes were evident in the roots of Al-treated plants.

Microsporidiosis in mammals.

Microsporidia are small, single-celled, obligately intracellular parasites that have caused significant agricultural losses and interference with biomedical research. Interest in the microsporidia is growing, as these organisms are recognized as agents of opportunistic infections in persons with AIDS and in organ transplant recipients. Microsporidiosis is also being recognized in children and travelers, and furthermore, concern exists about the potential of zoonotic and waterborne transmission of microsporidia to humans. This article reviews the basic biology and epidemiology of microsporidiosis in mammals.

Biology of microsporidian species infecting mammals.

Microsporidia (phylum Microspora) are obligate intracellular protozoan parasites that infect a wide range of vertebrate and invertebrate hosts. Over 1000 species have been classified into approximately 100 genera, and at least 13 species have been reported to infect mammals. Phylogenetically, the microsporidia are early eukaryotes because they have a true nucleus, possess prokaryote-like ribosomes, and lack mitochondria. The species that infect mammals are relatively small, measuring 2.0-7.0 microns long and 1.5-5.0 microns wide. The mature organism is the spore, which is enclosed by a chitinous coat, making it relatively resistant to the environment. Infections often occur by fecal-oral or urinary-oral transmission, although vertical transmission is quite common in the carnivores. Host cells become infected through a process of germination in which the spore propels its contents through the everting and unwinding polar filament into the host cell. The polar filament is unique to the microsporidia. With a few exceptions, microsporidiosis is typically chronic and subclinical in immunologically competent hosts. Young carnivores infected with microsporidia, however, develop severe and sometimes lethal renal disease, and immunodeficient laboratory animals (e.g. athymic and SCID mice) develop ascites and die from microsporidiosis. This review describes the morphology, life cycle, taxonomy, and host-parasite relationships of the species of microsporidia that infect mammals.

Antigen detection immunoassay using dipsticks and colloidal dyes.

A dipstick colloidal dye immunoassay (DIA) for multiple antigen detection is described. The test combines the concepts of double antibody (Ab) sandwich ELISA, dot blotting, and colloidal particle-linked Abs to produce a dipstick test for multiple antigen (Ag) detection. Dipsticks prepared from Ab coated nitrocellulose membrane mounted on acetate strips served as the assay capture matrix. Abs absorbed to colored dye particles from a family of commercially available textile dyes (Dye/Ab reagent) served as Ag detecting reagents. DIA and enzyme labelled dot blot assays showed similar Ag detection limits down to a sensitivity of 10 ng/ml. In a pilot study, an assay designed to detect species-specific IgG for use in mosquito bloodmeal identification demonstrated the feasibility of the technique. Experiments comparing bloodmeal analysis of mosquitoes using DIA and ELISA methods showed 100% agreement. This DIA method provides an inexpensive, simple, robust test for multiple Ag detection without instrumentation suitable for a wide variety of field applications.

Dynamics of immune responses related to clinical status in Brugia pahangi-infected dogs.

Clinical signs of lymph node enlargement, limb edema, lymph duct fibrosis, and microfilaremia were monitored in dogs with chronic Brugia pahangi infections. During the study a single rear limb of each dog was reinfected with multiple low doses of infective larvae. The changing immune responses to parasite antigens prepared from three sources--Brugia pahangi adult worm homogenate extract, adult worm excretory-secretory products, and microfilaria excretory-secretory products--were monitored by Western blot ELISA of antigens fractionated on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and by microtiter plate ELISA. Assays were used to detect antibodies in both the the IgG and IgE classes. A wide range of clinical manifestations was demonstrated in response to reinfection: asymptomatic, amicrofilaremic; asymptomatic, microfilaremic; acute short duration node enlargement and/or limb edema with microfilaremia; and chronic limb edema, amicrofilaremic. On microtiter plate ELISA, the dogs demonstrating the highest anti-adult worm homogenate titers were amicrofilaremic and were asymptomatic or developed chronic limb edema, dogs with high anti-mf ES titers were persistently amicrofilaremic, and the most marked increases against all three antigen sources upon reinfection occurred in low or amicrofilaremic dogs. Quantitative changes in antibody levels against the three crude antigen sources following reinfection were often paralleled by distinct changes in recognition of specific bands of antigens fractionated by SDS-PAGE.

Selective breeding of dogs for segregation of limb edema from microfilaremia as clinical manifestations of Brugia infections.

Three generations of beagles were monitored for microfilaremia (mf) and clinical disease during repeated infection with Brugia pahangi and were selectively bred for offspring manifesting limb edema and low or amicrofilaremia. A high microfilaremic female mated to a high microfilaremic male produced 7 pups, 6 of which maintained mf greater than 1,000/ml for greater than 2 years after 5 monthly infections of 10 infective larvae each. An uninfected female mated to another high mf male produced 5 pups, 4 of which did not exceed 1,000 mf/ml 7 months after initiation of the repeating infection regimen; 1 of these remained amicrofilaremic after 2 additional challenges. Neither the parents nor the offspring from these matings manifested chronic limb edema. Two matings were conducted with offspring from the microfilaremic female by breeding siblings with the lowest mf and breeding siblings with the highest mf. The high mf siblings produced 4/5 offspring manifesting chronic limb edema (greater than or equal to 7 months duration) and either no mf (in 2 dogs) or less than 100 mf/ml after the repeating infection regimen. The lower mf siblings produced 5 offspring, all with greater than 1,000 mf/ml 6 months after the initiation of the repeating infection regimen; none manifested edema. Comparisons of IgG antibody levels, specific for extracts of adult worms, showed no consistent differences between these 2 litters of dogs that could be associated with limb edema or mf when monitored for 16 months; however, the onset of lymph node enlargement was much earlier in the group of dogs manifesting limb edema than in the other litter.

Human cystic echinococcosis in a Uruguayan community: a sonographic, serologic, and epidemiologic study.

A prevalence and transmission study of human cystic echinococcosis (CE), due to infection with the dog tapeworm Echinococcus granulosus, was undertaken in the village of La Paloma in central Uruguay. The human population was registered and screened for CE by abdominal ultrasound scan as well as a number of serologic tests. Dogs were screened for E. granulosus infection by arecoline purgation as well as specific coproantigen testing. The total prevalence of human CE (new cases and those with a previous history) was 5.6% (64 of 1,149); 3.6% (40) of the cases were new ultrasound detected asymptomatic cases (mean age = 45 years). Age prevalence increased from 1.1% in the 4-6-year-old group to > 11% in the > 60-year-old group; the 20-29-year-old group had a significantly higher CE rate of 7.4%, compared with younger and older age groups, and there was no difference between sexes. A CE rate of 3.9% (20 of 514) was also recorded by ultrasound for new cases in the population residing outside the village. Most of the hydatid cysts were located in the liver presenting as either univesicular cysts or a solid mass, and of those 71% and 63%, respectively, with such cyst presentations were seropositive against E. granulosus cyst fluid antigens. Two of eight individuals who were filter paper blood spot seropositive, but ultrasound scan negative, were subsequently diagnosed respectively with pulmonary hydatidosis after radiography, and hepatic hydatidosis after computed tomography scan. Of 36 households with a CE patient, 32 were single cases while four households each harbored two CE cases. This did not represent a clustered distribution within families (23 of 117). Almost 20% of the dogs from La Paloma were found infected with E. granulosus after purge examination, with a mean worm number of 67 (range = 1-1,020). An additional eight dogs that were purge negative were Echinococcus coproantigen positive. The study showed that human CE is highly endemic in Uruguay, with one of the highest local prevalence rates in the world. Transmission appears to occur readily within well-developed towns, as well as on rural sheep ranches. Mass screening by ultrasound scanning with confirmatory serologic testing is an effective approach to case detection at the community level.

The lymphatic pathology of chronic Brugia pahangi infection in the dog.

The dog infected with Brugia pahangi is an excellent model for studying the clinical and immunological pathogenesis of lymphatic filarial infection and disease. Dogs demonstrated a range of clinical changes including varying levels of microfilaraemia, episodic lymphadenopathy, lymphangitis, and limb oedema similar to the clinical spectrum reported in man. Histologically, tissues from parasite-infected dogs were characterized by fibrosis of the lymph nodes and their afferent and efferent lymphatic ducts in the infected limbs. Some nodes were atrophied while others were characterized by reactive hyperplasia. Duct walls of some afferent lymphatics were thickened with a mononuclear inflammatory infiltrate. Parasite antigen deposition was detected in the subcapsular sinuses and middle of germinal centres of infected popliteal lymph nodes, using an immunohistochemical technique.