Accumulation, persistence and effects of indospicine residues in camels fed Indigofera plant

Indospicine (L-2-amino-6-amidinohexanoic acid) is a natural hepatotoxin found in all parts of some Indigofera plants such as I. linnaei and I. spicata. Several studies have documented a susceptibility to this hepatotoxin in different species of animals, including cattle, sheep, dogs and rats, which are associated with mild to severe liver disease after prolonged ingestion. However, there is little published data on the effects of this hepatotoxin in camels, even though Indigofera plants are known to be palatable to camels in central Australia. The secondary poisoning of dogs after prolonged dietary exposure to residual indospicine in camel muscle has raised additional food safety concerns. In this study, a feeding experiment was conducted to investigate the in vivo accumulation, excretion, distribution and histopathological effects of dietary indospicine on camels. Six young camels (2 – 4 year old), weighing 270 − 390 kg were fed daily a roughage diet consisting of Rhodes grass hay and lucerne chaff, supplemented with Indigofera and steam flaked barley. Indigofera (I. spicata) was offered at 597 mg DM/kg body weight (bw)/day designed to deliver 337 µg indospicine/kg bw/day, and fed for a period of 32 days. Blood and muscle biopsies were collected over the period of the study. Concentrations of indospicine in the plasma and muscle biopsy samples were quantitated by validated ultra-performance liquid chromatography−tandem mass spectrometry (UPLC−MS/MS). The highest concentrations in plasma (1.01 mg/L) and muscle (2.63 mg/kg fresh weight (fw)) were found at necropsy (day 33). Other tissues were also collected at necropsy and analysis showed ubiquitous distribution of indospicine, with the highest indospicine accumulation detected in the pancreas (4.86 ± 0.56 mg/kg fw) and liver (3.60 ± 1.34 mg/kg fw); followed by the muscle, heart and kidney. Histopathological examination of liver tissue showed multiple small foci of predominantly mononuclear inflammatory cells. After cessation of Indigofera intake, indospicine present in plasma in the remaining 3 camels had a longer terminal elimination half-life (18.6 days) than muscle (15.9 days), and both demonstrated mono-exponential decreases.

Nucleotide Degradation Profiles of Iced Barramundi (Lates calcarifer) and Nile Perch (Lates niloticus) Muscle

Adenine nucleotides and their related compounds were determined in muscle extracts from two species of fish that were stored in ice after thawing. The fish were the closely related species, Australian barramundi (Lates calcarifer ) and Kenyan Nile perch (Lates niloticus ) which had different process histories. For all samples, adenine nucleotides did not exceed 6% of the total nucleotide pool. Inosine monophosphate (IMP) decreased steadily with storage. Hypoxanthine (Hx) was the major product of adenosine triphosphate (ATP) degradation in both barramundi and Nile perch, showing a steady increase with days of iced storage. The Hx level did not reach a maximum during the 9d storage period. The K-value also increased regularly with time of storage and for the later stages (i.e., 7 and 9d) and was significantly different (P < 0.01) for the two species. The iced storage life of these typical samples of barramundi and Nile perch was estimated to be 3d after thawing using a K-value of < 30% to indicate excellent quality. Despite the differences in process history the nucleotide profiles were remarkably similar during storage. This precludes the use of nucleotide levels as a means of differentiating between these species.

Maternal and Paternal Genomes Differentially Affect Myofibre Characteristics and Muscle Weights of Bovine Fetuses at Midgestation

Postnatal myofibre characteristics and muscle mass are largely determined during fetal development and may be significantly affected by epigenetic parent-of-origin effects. However, data on such effects in prenatal muscle development that could help understand unexplained variation in postnatal muscle traits are lacking. In a bovine model we studied effects of distinct maternal and paternal genomes, fetal sex, and non-genetic maternal effects on fetal myofibre characteristics and muscle mass. Data from 73 fetuses (Day153, 54% term) of four genetic groups with purebred and reciprocal cross Angus and Brahman genetics were analyzed using general linear models. Parental genomes explained the greatest proportion of variation in myofibre size of Musculus semitendinosus (80-96%) and in absolute and relative weights of M. supraspinatus, M. longissimus dorsi, M. quadriceps femoris and M. semimembranosus (82-89% and 56-93%, respectively). Paternal genome in interaction with maternal genome (P<0.05) explained most genetic variation in cross sectional area (CSA) of fast myotubes (68%), while maternal genome alone explained most genetic variation in CSA of fast myofibres (93%, P<0.01). Furthermore, maternal genome independently (M. semimembranosus, 88%, P<0.0001) or in combination (M. supraspinatus, 82%; M. longissimus dorsi, 93%; M. quadriceps femoris, 86%) with nested maternal weight effect (5-6%, P<0.05), was the predominant source of variation for absolute muscle weights. Effects of paternal genome on muscle mass decreased from thoracic to pelvic limb and accounted for all (M. supraspinatus, 97%, P<0.0001) or most (M. longissimus dorsi, 69%, P<0.0001; M. quadriceps femoris, 54%, P<0.001) genetic variation in relative weights. An interaction between maternal and paternal genomes (P<0.01) and effects of maternal weight (P<0.05) on expression of H19, a master regulator of an imprinted gene network, and negative correlations between H19 expression and fetal muscle mass (P<0.001), suggested imprinted genes and miRNA interference as mechanisms for differential effects of maternal and paternal genomes on fetal muscle.

Real-time PCR as a surveillance tool for the detection of Trichinella infection in muscle samples from wildlife

Trichinella nematodes are the causative agent of trichinellosis, a meat-borne zoonosis acquired by consuming undercooked, infected meat. Although most human infections are sourced from the domestic environment, the majority of Trichinella parasites circulate in the natural environment in carnivorous and scavenging wildlife. Surveillance using reliable and accurate diagnostic tools to detect Trichinella parasites in wildlife hosts is necessary to evaluate the prevalence and risk of transmission from wildlife to humans. Real-time PCR assays have previously been developed for the detection of European Trichinella species in commercial pork and wild fox muscle samples. We have expanded on the use of real-time PCR in Trichinella detection by developing an improved extraction method and SYBR green assay that detects all known Trichinella species in muscle samples from a greater variety of wildlife. We simulated low-level Trichinella infections in wild pig, fox, saltwater crocodile, wild cat and a native Australian marsupial using Trichinella pseudospiralis or Trichinella papuae ethanol-fixed larvae. Trichinella-specific primers targeted a conserved region of the small subunit of the ribosomal RNA and were tested for specificity against host and other parasite genomic DNAs. The analytical sensitivity of the assay was at least 100 fg using pure genomic T. pseudospiralis DNA serially diluted in water. The diagnostic sensitivity of the assay was evaluated by spiking log of each host muscle with T. pseudospiralis or T. papuae larvae at representative infections of 1.0, 0.5 and 0.1 larvae per gram, and shown to detect larvae at the lowest infection rate. A field sample evaluation on naturally infected muscle samples of wild pigs and Tasmanian devils showed complete agreement with the EU reference artificial digestion method (k-value = 1.00). Positive amplification of mouse tissue experimentally infected with T. spiralis indicated the assay could also be used on encapsulated species in situ. This real-time PCR assay offers an alternative highly specific and sensitive diagnostic method for use in Trichinella wildlife surveillance and could be adapted to wildlife hosts of any region. (C) 2012 Elsevier B.V. All rights reserved.