Evidence for induction of humoral and cytotoxic immune responses against devil facial tumor disease cells in Tasmanian devils (Sarcophilus harrisii) immunized with killed cell preparations.

Tasmanian devils (Sarcophilus harrisii) risk extinction from a contagious cancer, devil facial tumour disease (DFTD) in which the infectious agent is the tumor cell itself. Because devils are unable to produce an immune response against the tumor cells no devil has survived 'infection'. To promote an immune response we immunized healthy devils with killed DFTD tumor cells in the presence of adjuvants. Immune responses, including cytotoxicity and antibody production, were detected in five of the six devils. The incorporation of adjuvants that act via toll like receptors may provide additional signals to break 'immunological ignorance'. One of these devils was protected against a challenge with viable DFTD cells. This was a short-term protection as re-challenge one year later resulted in tumor growth. These results suggest that Tasmanian devils can generate immune responses against DFTD cells. With further optimization of immune stimulation it should be possible to protect Tasmanian devils against DFTD with an injectable vaccine.

High phylogenetic diversity of the cat flea (Ctenocephalides felis) at two mitochondrial DNA markers.

The cat flea, Ctenocephalides felis (Siphonaptera: Pulicidae) (Bouché), is the most common flea species found on cats and dogs worldwide. We investigated the genetic identity of the cosmopolitan subspecies C. felis felis and evaluated diversity of cat fleas from Australia, Fiji, Thailand and Seychelles using mtDNA sequences from cytochrome c oxidase subunit I (cox1) and II (cox2) genes. Both cox1 and cox2 confirmed the high phylogenetic diversity and paraphyletic origin of C. felis felis. The African subspecies C. felis strongylus (Jordan) is nested within the paraphyletic C. felis felis. The south East Asian subspecies C. felis orientis (Jordan) is monophyletic and is supported by morphology. We confirm that Australian cat fleas belong to C. felis felis and show that in Australia they form two distinct phylogenetic clades, one common with fleas from Fiji. Using a barcoding approach, we recognize two putative species within C. felis (C. felis and C. orientis). Nucleotide diversity was higher in cox1 but COX2 outperformed COX1 in amino acid diversity. COX2 amino acid sequences resolve all phylogenetic clades and provide an additional phylogenetic signal. Both cox1 and cox2 resolved identical phylogeny and are suitable for population structure studies of Ctenocephalides species.

'Cysticercosis storm' in feedlot cattle in north-west New South Wales.

OBJECTIVE: To investigate the cause of an outbreak of bovine cysticercosis (Taenia saginata) infection on a cattle property in north-western New South Wales (NSW). METHODS: Cystic lesions were detected in the muscles of cattle during routine meat inspection at slaughter. These lesions were confirmed to be cysticerci of T. saginata through histology and polymerase chain reaction (PCR). Data on cattle maintenance were obtained through interviews with feedlot owners and staff. A suspect feed supplement was investigated. RESULTS: Between 5 July to 13 December 2010, 390 feedlot cattle from north-western NSW were slaughtered in abattoirs in NSW and Queensland. Of these, 138 animals had been maintained exclusively in feedlot enclosures from 80 to 300 days. Bovine cysticercosis was discovered in 80 cattle (58%, 26 carcasses were condemned). Another 18 cattle spent 24 h in the feedlot before being moved onto pasture and 1 of them was found to be infected. During the 5 months following the initial outbreak, a further 275 cattle were slaughtered; 2 of 51 heifers retained in the feedlot for a further 100 days were infected. None of the 234 animals grazed exclusively on pasture on the property were infected. Bovine cysticercosis was confirmed through examination of histological sections of muscle lesions and PCR using DNA extracted from cysticerci. No eggs of T. saginata were recovered from the feed supplement using a standard flotation method. CONCLUSIONS: The source of infection arose from rations contaminated with human faeces. All possibilities for local contamination were eliminated during the investigation. The suspected source of infection was imported copra meal, which was used as a feed supplement.

Changes in problem-solving appraisal after cognitive therapy for the prevention of suicide.

BACKGROUND: Cognitive therapy has been found to be effective in decreasing the recurrence of suicide attempts. A theoretical aim of cognitive therapy is to improve problem-solving skills so that suicide no longer remains the only available option. This study examined the differential rate of change in problem-solving appraisal following suicide attempts among individuals who participated in a randomized controlled trial for the prevention of suicide. METHOD: Changes in problem-solving appraisal from pre- to 6-months post-treatment in individuals with a recent suicide attempt, randomized to either cognitive therapy (n = 60) or a control condition (n = 60), were assessed by using the Social Problem-Solving Inventory-Revised, Short Form. RESULTS: Improvements in problem-solving appraisal were similarly observed for both groups within the 6-month follow-up. However, during this period, individuals assigned to the cognitive therapy condition demonstrated a significantly faster rate of improvement in negative problem orientation and impulsivity/carelessness. More specifically, individuals receiving cognitive therapy were significantly less likely to report a negative view toward life problems and impulsive/carelessness problem-solving style. CONCLUSIONS: Cognitive therapy for the prevention of suicide provides rapid changes within 6 months on negative problem orientation and impulsivity/carelessness problem-solving style. Given that individuals are at the greatest risk for suicide within 6 months of their last suicide attempt, the current study demonstrates that a brief cognitive intervention produces a rapid rate of improvement in two important domains of problem-solving appraisal during this sensitive period.

Reversible infantile respiratory chain deficiency is a unique, genetically heterogenous mitochondrial disease.

OBJECTIVES: Homoplasmic maternally inherited, m.14674T>C or m. 14674T>G mt-tRNA(Glu) mutations have recently been identified in reversible infantile cytochrome c oxidase deficiency (or 'benign COX deficiency'). This study sought other genetic defects that may give rise to similar presentations. PATIENTS: Eight patients from seven families with clinicopathological features of infantile reversible cytochrome c oxidase deficiency were investigated. METHODS: The study reviewed the diagnostic features and performed molecular genetic analyses of mitochondrial DNA and nuclear encoded candidate genes. RESULTS: Patients presented with subacute onset of profound hypotonia, feeding difficulties and lactic acidosis within the first months of life. Although recovery was remarkable, a mild myopathy persisted into adulthood. Histopathological findings in muscle included increased lipid and/or glycogen content, ragged-red and COX negative fibres. Biochemical studies suggested more generalised abnormalities than pure COX deficiency. Clinical improvement was reflected by normalisation of lactic acidosis and histopathological abnormalities. The m.14674T>C mt-tRNA(Glu) mutation was identified in four families, but none had the m. 14674T>G mutation. Furthermore, in two families pathogenic mutations were also found in the nuclear TRMU gene which has not previously been associated with this phenotype. In one family, the genetic aetiology still remains unknown. CONCLUSIONS: Benign COX deficiency is better described as 'reversible infantile respiratory chain deficiency'. It is genetically heterogeneous, and patients not carrying the m.14674T>C or T>G mt-tRNA(Glu) mutations may have mutations in the TRMU gene. Diagnosing this disorder at the molecular level is a significant advance for paediatric neurologists and intensive care paediatricians, enabling them to select children with an excellent prognosis for continuing respiratory support from those with severe mitochondrial presentation in infancy.

Two cases with progressive cystic leukoencephalopathy.

Leukoencephalopathies with cystic changes in the white matter on magnetic resonance imaging are aetiologically heterogeneous neurological disorders seen in children. A group of leukoencephalopathies characterised by white matter lesions progressing to multifocal cystic degeneration has been reported in various disorders, including mitochondrial enzyme deficiencies, leukodystrophies, and infectious processes. We report two patients with leukoencephalopathy showing progressive cystic changes on serial MRI, and magnetic resonance spectroscopy resembling progressive cavitating leukoencephalopathy.

A putative exonic splicing enhancer in exon 7 of the PDHA1 gene affects splicing of adjacent exons.

A nonsense mutation (c.729C>A, Y243X) in exon 7 of the PDHA1 gene in a patient with pyruvate dehydrogenase deficiency results in aberrant splicing of the primary transcript with production of stable mRNAs which lack either both exons 6 and 7 or exon 7 alone. Transfection and expression of genomic constructs covering exons 5 to 8 of the mutant PDHA1 gene reproduced this aberrant splicing in vitro. The same pattern of abnormal splicing was found when a silent mutation was introduced at the same position. Both the nonsense and silent mutations alter a strong consensus site for the binding of SRp40, suggesting that they may interfere with an exonic splicing enhancer in exon 7 of the gene. However, this appears to affect splicing of not only exon 7, but also the adjacent upstream exon. The splice acceptor site of intron 5 has weak homology to the consensus sequence and this may contribute to the combined splicing defect.

The link between depressive symptoms, negative cognitive bias and memory complaints in older adults.

OBJECTIVES: We sought to examine the relationship between depressive symptoms and subjective memory problems. We hypothesized that the relationship between depressive symptoms and poor subjective memory functioning is mediated by negative cognitive bias that is associated with hopelessness, a wish to die and low self-esteem. METHODS: Complete data were available for 299 older adults with and without significant depressive symptoms who were screened in primary care offices and invited to participate, completed a baseline in-home assessment. Subjective memory functioning and psychological status was assessed with commonly used, validated standard questionnaires. RESULTS: In regression models that included terms for age, gender and cognitive measures, depressive symptoms were significantly inversely associated with the global self-assessment of memory (beta=-0.019; p=0.006). When components of negative cognitive bias were included in the model (hopelessness, low self-esteem, a wish to die), the relationship of depressive symptoms with subjective memory problems was attenuated, consistent with mediation. CONCLUSIONS: Our results suggest that assessment and successful interventions for memory complaints in non-demented older adults need to account for negative cognitive bias as well as depressive symptoms. Longitudinal research is needed to confirm our findings before a mediator relationship can be presumed.

Mitochondrial cytopathies, phenotypic heterogeneity and a high incidence.

Mitochondrial respiratory chain disorders account for significant and varied presentations in paediatric practice. The true prevalence of these disorders in the paediatric population is still not well documented with predicted geographic variation. We report a retrospective analysis over a seven year period of cases presenting to a tertiary care centre and associated clinical features. The overall prevalence of mitochondrial disorders in our population is higher than expected (1/9,000 births), explained in part by multiple presentations in a consanguineous subgroup of the population (Irish travellers).

Detection of Anaplasma platys and Babesia canis vogeli and their impact on platelet numbers in free-roaming dogs associated with remote Aboriginal communities in Australia.

OBJECTIVE: To detect Anaplasma platys and Babesia canis vogeli infection, using polymerase chain reaction (PCR)-based assays, in free-roaming dogs associated with eight Aboriginal communities in remote areas of Australia and to determine the impact of infection through the assessment of platelet numbers. PROCEDURES: Blood samples from 215 dogs were screened by PCR for A platys and B canis vogeli using established genus-specific DNA primers for the 16S and 18S rRNA genes respectively. Both A platys DNA and B canis vogeli DNA were confirmed from the screening PCR either by sequencing or by the use of species-specific primers. Peripheral blood films from 92 of the 215 dogs were used to estimate platelet numbers through an indirect method. RESULTS: Of 215 dogs, 69 (32%) were positive for A platys, 22 (10%) for B canis vogeli and 24 (11%) for both. The two organisms were detected singularly and as coinfection in all communities. For the 92 dogs in which peripheral blood films were examined, the mean estimated platelet counts for the non-infected dogs was 318 x 10(9)/L, those infected with A platys alone was 256 x 10(9)/L, those with B canis vogeli alone was 276 x 10(9)/L and those infected with both parasites was 169 x 10(9)/L. In young dogs, infection produced significantly decreased mean platelet counts when compared to uninfected dogs. Thrombocytopenia (< 200 x 10(9)/L) was detected in 18 (51%) dogs infected with A platys alone, 3 (33%) dogs infected with B canis vogeli alone, 13 (72%) dogs coinfected, and 8 (27%) uninfected dogs. CONCLUSIONS: A platys and B canis vogeli infection, either singularly or together, was widespread in free roaming dogs associated with remote Aboriginal communities in the Northern Territory and north-western New South Wales. Moreover, both A platys and B canis vogeli infections were associated with a reduction in mean platelet numbers in dog populations, particularly in young dogs. The fact that 51% of dogs infected with A platys alone and 72% dogs coinfected were thrombocytopenic compared to 27% of uninfected dogs suggests that the organism alone or in combination with B canis vogeli has the potential to cause thrombocytopenia and perhaps contribute to a clinical bleeding disorder in infected dogs.

Pyruvate dehydrogenase E3 binding protein (protein X) deficiency.

Pyruvate dehydrogenase (PDH) deficiency is a major cause of neurological dysfunction and lactic acidosis in infancy and early childhood. The great majority of cases (>80%) result from mutations in the X-linked gene for the E1alpha subunit of the complex (PDHA1). Mutations in the genes for the other subunits have all been described, but only dihydrolipoamide dehydrogenase (E3) and E3 binding protein (E3BP) defects contribute significantly to the total number of patients with PDH deficiency. Although previously considered rare, with only 13 reported cases, we have found that mutations in PDX1, the gene for the E3 binding protein, are in fact relatively common. Clinical, biochemical, and genetic features of six new patients (four males, two females; age range 15mo-6y) with mutations in this gene are compared with previously reported cases. All patients with E3BP deficiency identified to date have mutations which completely prevent synthesis of the protein product. However, they are generally less severely affected than patients with PDHA1 mutations, although there is considerable overlap in clinical and neuroradiological features.

Congenital brain malformations in mitochondrial disease.

The majority of patients with mitochondrial disease have some degree of neuropathology and this is usually degenerative in nature. However, in a subset of mitochondrial diseases there are additional specific and characteristic congenital malformations in the brain. These developmental abnormalities are not unique to mitochondrial diseases and their association with only some conditions suggests that they are not simply due to energy deprivation or accumulation of metabolites such as lactic acid. Pathogenic mechanisms are at present unknown, but interference with neurotransmitter homeostasis, in addition to altered energy metabolism, may be important. The postnatal evolution of the neuropathology in these conditions and the type and pattern of the malformations suggest secondary degeneration of specific brain structures during fetal life rather than primary interference with developmental pathways.

Pyruvate dehydrogenase deficiency presenting as dystonia in childhood.

Two individuals with pyruvate dehydrogenase (PDH) deficiency due to missense mutations in the gene for the E1alpha subunit (PDHA1) presented during childhood with dystonia. The first patient, a male, presented at age 4 years with dystonia affecting the lower limbs, which responded to treatment with combined carbidopa and levodopa. The second patient, a female, was first investigated at age 6 years because of a dystonic gait disorder. In both patients, the main clue to the biochemical diagnosis was a raised concentration of lactate in the cerebrospinal fluid. PDH activity was significantly reduced in cultured fibroblasts in both cases. Dystonia is a previously unrecognized major manifestation of PDH deficiency and is of particular interest as the mutations in the PDHA1 gene in these patients have both been identified previously in individuals with typical presentations of the condition.

Marked increases in psychopathology found in a 30-year cohort comparison of suicide attempters.

BACKGROUND: Although several epidemiological studies have found increases in the percentages of people who have made a suicide attempt, few cohort comparisons have been conducted to determine changes within this population over time. The purpose of this investigation was to determine if there have been changes in the clinical profile of suicide attempters in recent decades. METHOD: Comparisons between a sample of 258 suicide attempters evaluated between 1970 and 1973 and a second sample of 179 suicide attempters evaluated between 1999 and 2002 were made on depression, hopelessness, suicide intent, drug use, history of suicide attempts and subsequent suicide attempts. RESULTS: Present-day suicide attempters were found to exhibit greater levels of depression (p = 0.031), hopelessness (p = 0.008), suicide intent (p < 0.001), and had much higher rates of illicit drug use (p < 0.001). Almost twice as many of the present-day suicide attempters had histories of four or more suicide attempts (p < 0.001), and the present-day suicide attempters made subsequent suicide attempts at close to four times the rate in the year following the index attempt (p < 0.001). CONCLUSIONS: The present-day suicide attempters exhibited greater levels of psychopathology on every major variable assessed. Replication is necessary and public health implications are discussed.

Lactic acidosis and developmental delay due to deficiency of E3 binding protein (protein X) of the pyruvate dehydrogenase complex.

Pyruvate dehydrogenase deficiency is an important cause of primary lactic acidosis. Most cases occur as a result of mutations in the gene for the E1 alpha subunit of the complex, with a small number resulting from mutations in genes for other components, most commonly the E3 and E3-binding protein subunits. We describe pyruvate dehydrogenase E3-binding protein deficiency in two siblings in each of two unrelated families from Kuwait. The index patient in each family had reduced pyruvate dehydrogenase activity in cultured fibroblasts and no detectable immunoreactive E3-binding protein. Both were homozygous for nonsense mutations in the E3-binding protein gene, one involving the codon for glutamine 266, the other the codon for tryptophan 5.

Diagnostic difficulties with common SURF1 mutations in patients with cytochrome oxidase-deficient Leigh syndrome.

In three unrelated patients with systemic cytochrome oxidase deficiency resulting from mutations in the SURF1 gene, the same mutation in the splice donor site of intron 3 was identified. All three patients were compound heterozygotes, two for the common insertion/deletion mutation in exon 4. In all three cases, complete definition of the causative mutations was only resolved by combined analysis of cDNA and genomic DNA. Several factors were identified that contributed to the diagnostic difficulties: preferential amplification of deleted cDNA, significant formation of heteroduplexes in cDNA PCR amplification and unequal representation of heterozygous peaks in genomic DNA sequences. These patients emphasize the need to perform mutation analysis on both cDNA and genomic DNA wherever possible.

A pathogenic glutamate-to-aspartate substitution (D296E) in the pyruvate dehydrogenase E1 subunit gene PDHA1.

In a patient with fatal neonatal lactic acidosis due to pyruvate dehydrogenase deficiency, the only potential mutation detected was c.888C>G in PDHA1, the gene for the E1alpha subunit of the complex. This would result in a substitution of glutamate for aspartate (D296E). Pathogenicity of this minor alteration in amino acid sequence was demonstrated by expression studies. By comparing the mutant sequence with the known structures of the E1 components of pyruvate dehydrogenase and the closely related branched chain alpha-ketoacid dehydrogenase, an explanation for the profound consequences of the mutation can be proposed.

Detection of Ehrlichia platys in dogs in Australia.

OBJECTIVE: To describe the detection of Ehrlichia platys in free-roaming dogs in Central Australia. PROCEDURE: Blood samples were collected from four dogs and examined for bacterial 16S ribosomal DNA using Polymerase Chain Reaction (PCR)-based assays. The three positive samples obtained were then sequenced and identification of the PCR product carried out. As a result of all three samples being identical to or closely related to part of the 16S rRNA gene of E. platys, blood samples were subsequently obtained from a further 24 dogs. These samples were screened using a PCR-assay to determine the presence of Ehrlichia DNA using genus-specific primers. The positive samples obtained from the screening process were then subjected to a further PCR-assay using E. platys specific primers. RESULTS: Of 28 dogs sampled, Ehrlichia DNA was detected in the blood of 13 dogs. Sequencing of the amplicons obtained indicated a high homology with the 16S rRNA gene for E. platys. When the E. platys-specific PCR was performed for 10 of those dogs, the 678 bp product obtained from the PCR amplification confirmed the identification as part of the 16S rRNA gene of E. platys in all 10 dogs. CONCLUSION: This study reports for the first time Ehrlichia carriage by dogs in Australia. It also indicates the usefulness of the PCR technique in rapidly and accurately identifying diseases that are otherwise difficult to detect. By using universal primers directed against bacterial 16S ribosomal DNA and sequencing analysis, the detection of potentially pathogenic Ehrlichia organisms that had not previously been found in Australia has been made possible.