Microbiological safety assessment of silkworm farms: a case study.

Silkworms have been farmed for their silk since ancient times. After silk reeling, their chrysalides are consumed as food in several Asian countries. Despite the long rearing tradition of this insect, few studies have investigated the silkworm's microbiological safety all along the life cycle, focusing on detecting silkworm pathogens or on the safety of the dried chrysalis for food consumption. However, the in-farm rearing process, which takes around forty days, may affect the microbial load of the silkworm and of the rearing environment, as well as the quality of fresh cocoon and other performance parameters. No data is available on how microbial contamination changes during the rearing period and between different farmers. Furthermore, in light of the possible use of the chrysalis as food, it is crucial to understand how its microbial load varies according to the water content. To address these specific questions, we conducted an investigation involving the analysis of specific microbial indicators commonly used in the food chain. We collected environmental and silkworm samples from several farms. The examination covered the entire life cycle of silkworms, beginning with the first instar larvae and concluding with the scrutiny of both freshly harvested and dried pupae. Silkworm farms in Northeast Italy proved to be an appropriate model system for carrying out the experimentation. Additionally, an evaluation of rearing performance was conducted, with a focus on the quality of fresh cocoons and the survival rate of the insects.

Food Safety Issues Related to Uses of Insects for Feeds and Foods.

Edible insects are expected to become an important nutrient source for animals and humans in the Western world in the near future. However, before insects can be put on the market, the safety of their use for feed and food is warranted. This literature study was prepared to provide an overview of the actual knowledge of possible food safety hazards, including chemical, microbiological, and allergenic agents and prions, to human and animal health upon the use of insects for food and feed, and to highlight data gaps and suggest the way forward. From the data available, heavy metals of concern are cadmium in black soldier fly and arsenic in yellow mealworm larvae. Investigated mycotoxins do not seem to accumulate. Residues of pesticides, veterinary drugs, and hormones, as well as dioxins and PCBs, are sometimes found in insects. Contamination of insects with pathogens to human health is a consequence of a combination of the substrates used and the farming and processing steps applied. Insects harbor a wide variety of microorganisms, and some human pathogenic bacteria may be present. In addition, insects may harbor and transmit parasites. There is no evidence so far insects may harbor pathogenic viruses or prions, but they may act as vectors. Insects and insect-derived products may have allergenic potential. In this review, evidence on some safety aspects is displayed, and data gaps are identified. Recommendations are given for future research to fill the most relevant data gaps.

Development of a clinical prediction score for detection of suspected cases of equine grass sickness (dysautonomia) in France.

Equine grass sickness (EGS) (equine dysautonomia) is a neurodegenerative condition of grazing equines. Pre-mortem diagnosis of EGS is a challenge for practitioners as definitive diagnosis requires ileal/myenteric lymph node biopsies. This study aimed to develop a clinical score that could be used by practitioners to improve the detection of acute or subacute EGS cases in the field. Suspected EGS cases were declared by veterinary practitioners. A case was classified as confirmed positive if ileal or rectal biopsy samples showed neuronal degeneration typical of EGS. A semi-quantitative scoring system, including epidemiological and clinical data, was created to attempt to classify suspected EGS horses into confirmed positive or negative cases. Each variable was weighted based on a boosted regression trees model, while taking into account its clinical relevance. Twenty-eight EGS cases were confirmed by biopsy during the entire study period. The best cut-off value for the score to have a high sensitivity while maximizing specificity was 8, with a sensitivity of 100% and a specificity of 53%. In our dataset, 77% of animals would be correctly classified with this cut-off value of 8. Highest sensitivity was chosen in order to detect the highest number of potential cases. Our score represents an inexpensive and useful tool to aid in the identification of suspected EGS cases in the field and selection for further diagnostics procedures to confirm or rule out the disease. Application of the score to larger populations of animals would be required to further adapt and refine the score.

Immunohistochemical Labelling for Cyclo-oxygenase-2: Does the Positive Control Guarantee Standardized Results?

Since the identification of cyclo-oxygenase-2 as a potentially important therapeutic target in veterinary oncology, numerous studies on its expression have been conducted. Unfortunately, results have been heterogeneous and conclusions are difficult to draw. We tested the ability of a defined positive control to guarantee reproducibility of results among different laboratories. Valid positive controls were defined by positivity of the renal macula densa without background labelling. Fifteen colorectal tumours and 15 oral squamous cell carcinomas were labelled immunohistochemically by six European laboratories. Slides were evaluated in blinded fashion for percentage of positive cells and labelling intensity by three pathologists, and results were analyzed statistically for reproducibility and inter-reader variability. Macula densa positivity was an insufficiently sensitive control to guarantee reproducible results for percentage of positive cells and labelling intensity. Inter-reader variability was proven statistically, making the case for image analysis or other automated quantitative evaluation techniques.

Digital squamous cell carcinoma in dogs: epidemiological, histological, and immunohistochemical study.

Squamous cell carcinoma represents 47.4% of all malignant canine digital lesions, but despite its frequency, there are few published studies available. Pathology submission records of 154 cases and follow-up of 49 animals were analyzed. On the 49 cases, histological evaluation was performed of the differentiation degree, mitotic index, presence of emboli, and immunohistochemical expression of vimentin and E-cadherin. The mean (SD) age of affected animals was 10.2 (2.3) years; no sex predisposition was recorded. Beauceron and Briard were 2 new overrepresented breeds. Dark-haired animals comprised 97 of 105 (92%); 94 dogs of 125 (75.2%) belonged to large and giant breeds. The forelimb was affected twice more than the hind limb. Probable metastases were observed in 4 dogs; new tumor development was recorded in 11 of 49 (22.4%). Epidemiologic factors, histological grade, mitotic index, and expression of immunohistochemical markers seemed not to be related to the clinical outcome.

Retrospective study of Mycoplasma gallisepticum meningoencephalitis in six turkey flocks in western France.

Mycoplasma gallisepticum (MG) meningoencephalitis was diagnosed in six turkey flocks, from 1998 to 2005, in the western part of France. Affected birds were 8-11 weeks old and all displayed neurological signs, especially torticollis, with more than half having concomitant respiratory signs. Microscopical examination of brain samples from birds in all six flocks revealed similar lesions of acute to subacute multifocal parenchymal necrosis, perivascular cuffing, leptomeningitis and vasculitis. Birds from four of the six affected flocks were seropositive for MG and in birds from four flocks MG DNA was detected by polymerase chain reaction performed on tracheal swabs or on samples of formalin-fixed and paraffin wax-embedded brain. To our knowledge, this is the first pathological description of naturally occurring cases of turkey MG meningoencephalitis in Europe.

Spinal cryptococcoma in an immunocompetent cat.

This report describes an unusual case of primary cryptococcoma in the proximal thoracic spinal cord of an 11-year-old immunocompetent cat from a farm on which there were large numbers of pigeons. This animal was referred for examination with progressive paralysis and shown to be free from feline immunodeficiency virus, feline leukaemia virus, feline coronavirus and Toxoplasma gondii. It died 2 months later. At necropsy, the only lesion detected was a malacic area, 4cm in length, in the spinal cord. Histopathological examination of the spinal cord revealed severe granulomatous inflammation associated with large numbers of encapsulated yeast cells. In addition to the granulomatous host response, necrosis, digestion chambers, Gitter cells, spheroids and lymphocytic perivascular cuffs were features of the malacic areas. Immunohistochemistry confirmed the presence of Cryptococcus neoformans var. grubii yeast cells.

Ovarian epithelioid trophoblastic tumor in a cynomolgus monkey.

Epithelioid trophoblastic tumor (ETT) is an unusual type of trophoblastic tumor, with features resembling carcinoma. In this study, we describe a 4-year-old cynomolgus monkey (Macaca fascicularis) showing, at necropsy, a lobulated mass replacing the left ovary and several nodular lesions within the lungs. Histologically, the mass in the ovary and lung metastases were characterized by nests of epithelioid cells, with intermingled, occasional, multinucleate tumor cells consistent with syncytiotrophoblasts and moderate amount of eosinophilic, hyaline-like material. Immunohistochemically, the tumor cells were diffusely positive for cytokeratins (AE1/AE3) and inhibin-alpha, but only focal immunoreactivity was observed for human chorionic gonadotropin, whereas placental alkaline phosphatase was always negative. On the basis of morphology and immunohistochemical reactivity, tumor cells were identified as intermediate trophoblast.

Antitumour and antiangiogenic effects of IDN 5390, a novel C-seco taxane, in a paclitaxel-resistant human ovarian tumour xenograft.

IDN 5390 is a novel C-seco taxane analogue selected for preclinical development on the basis of its antimotility activity on endothelial cells, antitumour efficacy in a large panel of human tumour xenografts and high tolerability in mouse. On the basis of oral availability, IDN 5390 is suitable for protracted administration schedules. Such a treatment schedule has been reported as the most appropriate to exploit the antiangiogenic effects of cytotoxic drugs. An ability to downregulate angiogenesis-related growth factors in tumour cells has been described for IDN 5390. The aim of the study was to investigate the antitumour and antiangiogenic potential of oral IDN 5390 on a human ovarian carcinoma xenograft, the INT.ACP/PTX, resistant to paclitaxel (PTX). Such tumour line was derived in vivo from a cisplatin-resistant tumour line, the A2780/DDP, which is sensitive to PTX. Compared to the parental cells, INT.ACP/PTX cells exhibited a high level of Pgp expression, resulting in a reduced in vitro sensitivity to both PTX and IDN 5390. The INT.ACP/PTX tumour xenograft was still resistant to PTX, but responsive to IDN 5390, when delivered per os, by a daily prolonged schedule. A direct effect on tumour cells, allowed by the high tolerability of the compound in mouse, cannot be excluded in vivo. Immunohistochemical analysis indicated a significant reduction of microvessel density in IDN 5390-treated tumours, lasting till 7 days after the last drug administration. Thus, a prolonged inhibitory effect on tumour angiogenesis is consistent with the persistent growth control of INT.ACP/PTX tumour achieved by IDN 5390. On the contrary, the low tolerability and the limited oral availability of conventional taxanes do not allow an easy feasibility of such treatment regimen. Thus, the tolerability profile of IDN 5390 in preclinical systems and its efficacy in PTX-resistant tumours support the therapeutic interest for its clinical development, with particular attention to oral daily prolonged schedules.

Slow-like electrostimulation switches on slow myosin in denervated fast muscle.

Adult fast and slow skeletal muscles are composed of a large number of fibers with different physiological and biochemical properties that under neuronal control can respond in a plastic manner to a variety of stimuli. Although muscle cells synthesize muscle-specific contractile proteins in the absence of motoneurons, after innervation the neuron controls the particular set of isoforms subsequently synthesized. However, agreement has not been reached on the mechanism, either chemotrophic or impulse-mediated, by which the nerve influences gene expression in the muscle. Here we report the effect on isomyosins of continuous, low-frequency (a protocol mimicking the discharge pattern of the slow motoneuron) direct electrical stimulation of a permanently denervated fast muscle, the extensor digitorum longus of adult rat. After several weeks, unlike sham-stimulated muscle, the stimulated muscle showed a dramatic increase of the slow myosin light and heavy chains. Myosin light chains were identified by two-dimensional gel electrophoresis. The slow myosin heavy chain was clearly distinguished from fast and embryonic types by one-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and orthogonal peptide mapping. The myosin change could be restricted to a portion of the muscle by the position of the stimulating electrodes. Taking into account the morphologic appearance of the electrostimulated muscle and the large body of evidence demonstrating the absolute dependence of slow myosin on specific innervation, our observations indicate that at least the slow motoneuron influences the isomyosin genes' expression by the kind of activity it imposes on developing muscle fibers.