PubMed Health⌕ Search

Biomedical subjects

M T Long

Publications and source records attributed to M T Long.

At least 19 recordsLinked to original sources
PubMed2004

Leukoencephalitis associated with selective viral replication in the brain of a pony with experimental chronic equine infectious anemia virus infection.

Neurologic disease occurs sporadically in horses infected with the equine infectious anemia virus (EIAV). This report describes a case of clinically severe neurologic disease in a pony experimentally infected with EIAV. This pony did not have fever or anemia, which are the characteristic clinical signs of disease. The histopathologic changes were characterized as lymphohistiocytic periventricular leukoencephalitis. Polymerase chain reaction and in situ hybridization data showed that the brain lesions were directly associated with viral replication and that high-level viral replication occurred selectively within the lesion and not in other tissues. These findings suggest that EIAV-associated neurologic disease is the direct result of viral replication.

Animals↗
PubMed2002

Fate of patulin in the presence of the yeast Saccharomyces cerevisiae.

Patulin is known to become analytically non-detectable during the production of cider from contaminated apple juice. The fate of [14C]-labelled patulin during the alcoholic fermentation of apple juice was studied. Three commercial cider strains of Saccharomyces cerevisiae degraded patulin during active fermentative growth, but not when growing aerobically. The products of patulin degradation were more polar than patulin itself and remained in the clarified fermented cider. Patulin did not appear to bind to yeast cells or apple juice sediment in these model experiments. HPLC analysis of patulin-spiked fermentations showed the appearance of two major metabolites, one of which corresponded by both TLC and HPLC to E-ascladiol prepared by the chemical reduction of patulin using sodium borohydride. Using a diode array detector, both metabolites had a lambda(max) = 271 nm, identical to that of ascladiol. The nmr spectrum of a crude preparation of these metabolites showed signals corresponding to those of the E-ascladiol prepared chemically and a weaker set of signals corresponding to those reported in the literature for Z-ascladiol.

Alcoholic Beverages↗
PubMed2000

Neutralization of maternal IL-4 modulates congenital protozoal transmission: comparison of innate versus acquired immune responses.

IL-4 levels were modulated in mice to test the hypothesis that induction of a maternal type 1 response would decrease the frequency of congenital Neospora caninum transmission. This hypothesis tested the relationship between IL-4 and both innate and adaptive immunity utilizing two basic experimental designs. In the first, maternal IL-4 was neutralized with mAb during pregnancy in naive mice concomitant with initial, virulent infection. In the second, maternal IL-4 was neutralized before pregnancy concomitant with a priming inoculation consisting of live, avirulent N. caninum tachyzoites followed by virulent challenge during subsequent gestation. In mice that were naive before pregnancy, neutralization of IL-4 during gestational challenge did not result in decreased congenital transmission as measured by PCR performed on 1-day-old neonatal mice. In mice that were primed and modulated before pregnancy, congenital transmission from gestational challenge was significantly decreased compared with control mice. Reduction in transmission constituted a decrease in the numbers of mice transmitting N. caninum and a lower frequency of transmission by individual dams (p < 0.05). Decreased congenital transmission was associated with significantly lower levels of maternal splenocyte IL-4 secretion, lower IL-4 mRNA levels, and higher levels of IFN-gamma secretion. Protected mice had significantly decreased Neospora-specific IgG1 compared with nonmodulated mice. These studies define a relationship between maternal Ag-specific immunity and the frequency of congenital transmission and demonstrate that modulation of type 2 cytokine responses can change the frequency of congenital protozoal transmission.

Animals↗
PubMed1999

Interferon-gamma and interleukin-12 mediate protection to acute Neospora caninum infection in BALB/c mice.

The type of immune response required to protect mice against clinical disease during acute Neospora caninum challenge was investigated in BALB/c mice. Groups of female BALB/c mice were infected i.p. with N. caninum tachyzoites concomitant with either: (1) antibody to interferon-gamma; (2) recombinant murine interleukin-12; or (3) recombinant murine interleukin-12 plus antibody to interferon-gamma. Mice treated with anti-interferon-gamma alone had increased morbidity/mortality, decreased body weight, increased foci of liver necrosis and increased numbers of N. caninum tachyzoites in the lung by 7 days p.i. compared with controls. Increased disease and parasite load in the anti-interferon-gamma-treated mice was associated with antigen-specific antibody IgG1 > IgG2a and a three-fold decreased ratio of antigen-specific interferon-gamma:interleukin-4. Mice treated with recombinant murine interleukin-12 had decreased encephalitis and brain parasite load at 3 weeks p.i. compared with control mice treated with PBS. In recombinant murine interleukin-12-treated mice, decreased brain lesions and parasite load were associated with antigen-specific antibody IgG2a > IgG1 and a three-fold increased ratio of antigen-specific interferon-gamma:interleukin-4 from splenocytes; the interleukin-12 effect was dependent upon interferon-gamma, as indicated by concomitant in vivo interferon-gamma neutralisation. By 6 weeks p.i. with N. caninum, there were no differences in brain lesions and parasite load between interleukin-12- and PBS-treated groups, indicating that the effects of interleukin-12 on driving a protective type 1 response were transient. These data indicate a role for interferon-gamma, interleukin-12 and type 1 immune responses in control of acute neosporosis in mice.

Animals↗
PubMed1999

Detection by PCR of Neospora caninum in fetal tissues from spontaneous bovine abortions.

The routine diagnosis of Neospora caninum abortion is based upon histopathologic changes in fetal tissues and identification of tissue parasites by immunohistochemistry. Confirmation of N. caninum infection by immunohistochemistry has low sensitivity. In the present study, we examined the utility of PCR in detecting N. caninum infection in fetal tissues from spontaneous bovine abortion. DNA was obtained from fresh and formalin-fixed tissues from 61 bovine fetuses submitted for abortion diagnosis. Histopathology and immunohistochemistry determined the true status of N. caninum infection in each fetus. In formalin-fixed paraffin-embedded tissues, PCR detected N. caninum DNA in 13 of 13 true-positive fetuses (100%) and in 1 of 16 true-negative fetuses (6%). In fresh or frozen tissues, PCR detected N. caninum DNA in 10 of 13 true-positive fetuses (77%) and 0 of 11 true-negative fetuses (0%). PCR also detected N. caninum DNA in 6 of 8 fetuses that had typical lesions of N. caninum but were immunohistochemistry negative, indicating a higher sensitivity of PCR in comparison to that of immunohistochemistry. N. caninum DNA was amplified most consistently from brain tissue. PCR detection of N. caninum DNA in formalin-fixed, paraffin-embedded tissues was superior to that in fresh tissues, presumably because of the increased accuracy of sample selection inherent in histologic specimens.

Abortion, Veterinary↗
PubMed1998

Comparison of intracerebral parasite load, lesion development, and systemic cytokines in mouse strains infected with Neospora caninum.

Neospora caninum, an apicomplexan parasite closely related to Toxoplasma gondii, causes abortion, stillbirths, and congenital neurologic disease in multiple animal species. The present study focuses on the development of encephalitis and intracerebral parasite load that occurs 6 wk postinfection (PI). Utilizing BALB/c, C57BL/6, and B10.D2 mice, an initial investigation was undertaken to determine the relative resistance of inbred strains to N. caninum-induced encephalitis. Relative resistance was defined in terms of central nervous system lesion development and parasite load. Based on other protozoal infections in mice, it was hypothesized that BALB/c and C57BL/6 should be contrasting in their relative resistance to N. caninum, with BALB/c and congenic B10.D2 mice less susceptible than C57BL/6 mice. Contrary to expectation, BALB/c and C57BL/6 were both highly susceptible to the development of N. caninum-induced encephalitis, whereas B10.D2 mice were resistant. Both BALB/c mice and C57BL/6 mice had significantly higher numbers of brain lesions and intracerebral tachyzoites than B10.D2 mice. Resistance in B10.D2 was associated with a high interferon (IFN)-gamma: interleukin (IL)-4 ratio from antigen-stimulated splenocytes, whereas susceptibility in C57BL/6 and BALB/c mice corresponded with a low splenocyte IFN-gamma: IL-4 ratio. In vivo measurement of Neospora-specific isotype antibodies demonstrated predominately IgG2a in serum from B10.D2 mice and IgG1 in serum from BALB/c and C57BL/6 mice. In conclusion, susceptibility of mice to N. caninum is unique compared to other protozoal diseases. The present study also demonstrates that parasite load is a fundamental measurement for evaluating disease induced by N. caninum and that a type 1 cytokine response may be necessary for regulation of this parameter.

Animals↗
PubMed1997

Enzymology of oxidation of tropic acid to phenylacetic acid in metabolism of atropine by Pseudomonas sp. strain AT3.

Pseudomonas sp. strain AT3 grew with dl-tropic acid, the aromatic component of the alkaloid atropine, as the sole source of carbon and energy. Tropic acid-grown cells rapidly oxidized the growth substrate, phenylacetaldehyde, and phenylacetic acid. Crude cell extracts, prepared from dl-tropic acid-grown cells, contained two NAD+-linked dehydrogenases which were separated by ion-exchange chromatography and shown to be specific for their respective substrates, dl-tropic acid and phenylacetaldehyde. Phenylacetaldehyde dehydrogenase was relatively unstable. The stable tropic acid dehydrogenase was purified to homogeneity by a combination of ion-exchange, molecular-sieve, and affinity chromatography. It had a pH optimum of 9.5 and was equally active with both enantiomers of tropic acid, and at this pH, phenylacetaldehyde was the only detectable product of tropic acid oxidation. The formation of phenylacetaldehyde from tropic acid requires, in addition to dehydrogenation, a decarboxylation step. By analogy with NAD+-specific isocitrate and malate dehydrogenases, phenylmalonic semialdehyde, a 3-oxoacid, would be expected to be the precursor of phenylacetaldehyde. Other workers have established that isocitrate and malate dehydrogenases catalyze the decarboxylation of enzyme-bound or added 3-oxoacid intermediates, a reaction that requires Mn2+ or Mg2+ ions. Studies with tropic acid dehydrogenase were hampered by lack of availability of phenylmalonic semialdehyde, but in the absence of added divalent metal ions, both enantiomers of tropic acid were completely oxidized and we have not, by a number of approaches, found any evidence for the transient accumulation of phenylmalonic semialdehyde.

Acetaldehyde↗
PubMed1996

Fetal loss in BALB/C mice infected with Neospora caninum.

The suitability of mice as a model for reproductive loss due to Neospora caninum infection was investigated. Groups of mice were infected with 2 x 10(6) N. caninum before pregnancy (10 days) and during pregnancy (days 5 and 10 of gestation). In mice infected before and during early pregnancy, fetal loss was evaluated throughout gestation, and pregnancy loss was evaluated by enumeration of fetal resorptions and total fetuses. In mice infected before pregnancy, no difference was present in resorptions between infected and control mice, although litter size was decreased in the infected mice (P < 0.05). In mice infected during early pregnancy (day 5 gestation) and examined temporally throughout gestation, resorptions were increased in the infected mice compared to the control mice (P < 0.05). In mice infected at 5 days gestation and examined at one time point (day 14 of gestation), the resorption rate for infected mice was 33% and 12% for controls (P < 0.05). Routine histopathologic examination and immunohistochemistry using N. caninum-specific antisera did not identify tachyzoites in placental and fetal tissues during the pre- and early pregnancy infections. In mice infected late midgestation (day 10), N. caninum tachyzoites were identified in placenta and fetal muscle and neural tissue. In the placenta, there was multifocal necrosis and hemorrhage with intralesional tachyzoites. Tachyzoites in fetal tissues were not associated with pathologic changes. No reproductive loss was associated with mice infected late in gestation. These data demonstrate that mice can be used as a model for the study of fetal resorption and congenital infection associated with N. caninum.

Animals↗
PubMed1995

Tropine dehydrogenase: purification, some properties and an evaluation of its role in the bacterial metabolism of tropine.

Tropine dehydrogenase was induced by growth of Pseudomonas AT3 on atropine, tropine or tropinone. It was NADP(+)-dependent and gave no activity with NAD+. The enzyme was very unstable but a rapid purification procedure using affinity chromatography that gave highly purified enzyme was developed. The enzyme gave a single band on isoelectric focusing with an isoelectric point at approximately pH 4. The native enzyme had an M(r) of 58,000 by gel filtration and 28,000 by SDS/PAGE and therefore consists of two subunits of equal size. The enzyme displayed a narrow range of specificity and was active with tropine and nortropine but not with pseudotropine, pseudonortropine, or a number of related compounds. The apparent Kms were 6.06 microM for tropine and 73.4 microM for nortropine with the specificity constant (Vmax/Km) for tropine 7.8 times that for pseudotropine. The apparent Km for NADP+ was 48 microM. The deuterium of [3-2H]tropine and [3-2H]pseudotropine was retained when these compounds were converted into 6-hydroxycyclohepta-1,4-dione, an intermediate in tropine catabolism, showing that the tropine dehydrogenase, although induced by growth on tropine, is not involved in the catabolic pathway for this compound. 6-Hydroxycyclohepta-1,4-dione was also implicated as an intermediate in the pathways for pseudotropine and tropinone catabolism.

Alcohol Oxidoreductases↗
PubMed1995

Biological production of semisynthetic opiates using genetically engineered bacteria.

Semisynthetic derivatives of morphine and related alkaloids are in widespread clinical use. Due to the complexity of these molecules, however, chemical transformations are difficult to achieve in high yields. We recently identified the powerful analgesic hydromorphone as an intermediate in the metabolism of morphine by Pseudomonas putida M10. Here we describe the construction of recombinant strains of Escherichia coli that express morphine dehydrogenase and morphinone reductase. These strains are capable of efficiently transforming the naturally occurring alkaloids morphine and codeine to hydromorphone and the antitussive hydrocodone, respectively. Our results demonstrate the potential for recombinant DNA technology to provide biological routes for the synthesis of known and novel semisynthetic opiate drugs.

Alcohol Oxidoreductases↗
PubMed1995

Transformations of morphine alkaloids by Pseudomonas putida M10.

The oxidation of morphine by washed-cell incubations of Pseudomonas putida M10 gave rise to a large number of transformation products including hydromorphone (dihydromorphinone), 14 beta-hydroxymorphine, 14 beta-hydroxymorphinone, and dihydromorphine. Similarly, in incubations with oxymorphone (14 beta-hydroxydihydromorphinone) as substrate, the major transformation product was identified as oxymorphol (14 beta-hydroxydihydromorphine). The identities of all these biological products were confirmed by mass spectrometry and 1H nuclear magnetic resonance spectroscopy. This is the first report describing structural evidence for the biological synthesis of 14 beta-hydroxymorphine and 14 beta-hydroxymorphinone. These products have applications as intermediates in the synthesis of semisynthetic opiate drugs.

Analgesics, Opioid↗
PubMed1995

Identification of Ehrlichia risticii as the causative agent of two equine abortions following natural maternal infection.

Two pregnant mares diagnosed as having equine monocytic ehrlichiosis based on history, clinical signs, and high serum antibody titers to Ehrlichia risticii aborted subsequent to recovery from illness. Mare 1 and mare 2 experienced clinical illness at 120 and 143 days of gestation and aborted at 203 and 226 days of gestation, respectively. The fetuses were expelled in fresh condition, and both mares retained their placentas upon abortion. Gross findings for the fetuses included meconium staining and petechiation of external surfaces. Internally, there was increased volume of feces within the small and large intestines and liver discoloration with enlargement. Microscopic findings included lymphohistiocytic enterocolitis, hepatitis, and myocarditis. Lymphoid hyperplasia and depletion were present in spleen, thymus, and lymph nodes. Ehrlichia risticii was recovered from bone marrow, spleen, lymph node, colon, and liver of the first fetus and bone marrow and colon of the second fetus. Electron microscopic evaluation of the organism isolated in cell culture revealed morphology consistent with E. risticii. The isolated organism was inoculated into a naive pony, and this pony developed high levels of antibody against E. risticii, became ehrlichemic, and developed clinical signs of depression, anorexia, and mild diarrhea. These findings confirm that E. risticii is an abortifacient under conditions of natural infection and should be considered as a differential diagnosis of equine abortions.

Abortion, Septic↗
PubMed1995

Evaluation of fetal infection and abortion in pregnant ponies experimentally infected with Ehrlichia risticii.

Fetal infectivity of Ehrlichia risticii was investigated in 19 ponies that were E risticii negative on the basis of results of an indirect fluorescent antibody (IFA) test. Thirteen pregnant ponies were infected by IV administration of E risticii between 90 and 180 days of gestation. Six pregnant ponies served as noninfected controls. Each infected pony had clinical signs of equine monocytic ehrlichiosis, was confirmed to be ehrlichemic, and developed an IFA titer to E risticii. Two infected ponies became recumbent, were unresponsive to supportive care, and were euthanatized. After recovery from clinical illness, the remaining ponies were observed throughout gestation for reproductive abnormalities. On abortion, each fetus was necropsied and tissue specimens from the liver, bone marrow, spleen, colon, and mesenteric lymph nodes were inoculated into canine monocyte cell cultures. Six infected ponies aborted at a mean 217 days of gestation, which was between postinoculation days 65 and 111. Five fetuses were recovered for evaluation, and E risticii was isolated from 4 of them. All 5 fetuses recovered had similar histologic finding, including enterocolitis, periportal hepatitis, and lymphoid hyperplasia with necrosis of the mesenteric lymph nodes and spleen. All 5 fetuses tested negative for IgG to E risticii, although 3 had low IgM titer to E risticii. The remaining 5 infected ponies had normal parturition. Presuckle IFA titer to E risticii was measured in 4 of the term foals, and results for 3 were positive. Two foals from infected ponies were monitored for 6 months and daily gain in body weight was comparable to that of a control foal. None of the control ponies became ill or seroconverted during the clinical illness phase, and none aborted throughout gestation Two control ponies seroconverted to E risticii 6 weeks before parturition. Results of this study indicate that E ristcii is a primary abortifacient under experimental conditions.

Abortion, Veterinary↗
PubMed1994

Cultural, molecular, and immunological characterization of the etiologic agent for atypical canine ehrlichiosis.

More than 100 cases of canine ehrlichiosis, with three fatalities, were serologically negative by the indirect immunofluorescent antibody (IFA) test with Ehrlichia canis or E. sennetsu antigen but were reactive at titers of 10 to 640 with E. risticii. Ehrlichia-like agents were isolated from three such cases. The agents isolated from those cases were morphologically indistinguishable from each other and from a prototype, E. risticii, the etiologic agent of equine monocytic ehrlichiosis, in terms of growth characteristics and by light or electron microscopy. The patterns of and products from PCR were identical to those of E. risticii. The 16S rRNA sequences were distinct from those of E. canis and E. ewingii but were identical to those of E. risticii. A PCR product corresponding to the 5' half of the 16S rRNA gene was obtained from amplification of DNA from E. risticii and both sources of the atypical canine ehrlichiosis agent but was not obtained from uninfected host cells. The entire sequence of 719 nucleotides was identical for all three sources. The percentages of relatedness of the partial 16S rRNA gene of the atypical canine ehrlichiosis agent to E. risticii, E. sennetsu, E. platys, E. equi, E. phagocytophila, E. canis, E. chaffeensis, and E. ewingii were 100.0, 98.9, 83.7, 83.0, 83.0, 82.2, 81.8, and 81.5, respectively. These data are consistent with the identity of these isolates as E. risticii. The caninotropic characteristics of naturally acquired infections due to E. risticii are herein described for the first time, and the epizootiological implications are discussed in relation to the host range of E. risticii, which may include dogs as reservoirs.

Animals↗
PubMed1993

The isolation and identification of 6-hydroxycyclohepta-1,4-dione as a novel intermediate in the bacterial degradation of atropine.

Growth of Pseudomonas AT3 on the alkaloid atropine as its sole source of carbon and nitrogen is nitrogen-limited and proceeds by degradation of the tropic acid part of the molecule, with the metabolism of the tropine being limited to the point of release of its nitrogen. A nitrogen-free compound accumulated in the growth medium and was isolated and identified as 6-hydroxycyclohepta-1,4-dione. This novel compound is proposed as an intermediate in tropine metabolism. It served as a growth substrate for the organism and was also the substrate for an NAD(+)-linked dehydrogenase present in cell extracts. The enzyme was induced during the tropine phase of diauxic growth on atropine or during growth on tropine alone.

Alcohol Oxidoreductases↗