Comparative Evaluation of Molecular Diagnostic Tests for Nucleospora salmonis and Prevalence in Migrating Juvenile Salmonids from the Snake River,USA

Abstract

Nucleospora salmonis is an intranuclear microsporidian that primarily infects lymphoblast cells and contributes to chronic lymphoblastosis and a leukemia-like condition in a range of salmonid species. The primary goal of this study was to evaluate the prevalence of N. salmonis in out-migrating juvenile hatchery and wild Chinook salmon Oncorhynchus tshawytscha and steelhead O. mykiss from the Snake River in the U.S. Pacific Northwest. To achieve this goal, we first addressed the following concerns about current molecular diagnostic tests for N. salmonis: (1) nonspecific amplification patterns by the published nested polymerase chain reaction (nPCR) test, (2) incomplete validation of the published quantitative PCR (qPCR) test, and (3) whether N. salmonis can be detected reliably from nonlethal samples. Here, we present an optimized nPCR protocol that eliminates nonspecific amplification. During validation of the published qPCR test, our laboratory developed a second qPCR test that targeted a different gene sequence and used different probe chemistry for comparison purposes. We simultaneously evaluated the two different qPCR tests for N. salmonis and found that both assays were highly specific, sensitive, and repeatable. The nPCR and qPCR tests had good overall concordance when DNA samples derived from both apparently healthy and clinically diseased hatchery rainbow trout were tested. Finally, we demonstrated that gill snips were a suitable tissue for nonlethal detection of N. salmonis DNA in juvenile salmonids. Monitoring of juvenile salmonid fish in the Snake River over a 3-year period revealed low prevalence of N. salmonis in hatchery and wild Chinook salmon and wild steelhead but significantly higher prevalence in hatchery-derived steelhead. Routine monitoring of N. salmonis is not performed for all hatchery steelhead populations. At present, the possible contribution of this pathogen to delayed mortality of steelhead has not been determined.

Received September 14, 2010; accepted November 14, 2010     

Farmers' perceptions and practices for managing tomato leaf curl virus disease in southern India

A survey of 174 tomato farmers in five districts of Karnataka State, South India, was conducted between March and July 1998 with the objectives of assessing farmers' knowledge and perceptions of tomato leaf curl virus disease (ToLCVD) and examining their current control practices. The findings showed that farmers were familiar with ToLCVD and regarded it as a serious constraint upon production, particularly in the hot season. However, only 2% of farmers were aware that tomato leaf curl virus (ToLCV) was transmitted by a whitefly vector, Bemisia tabaci . A similarly small number of farmers knew that ToLCVD was caused by a virus, whilst 86% of farmers believed that it was caused by high temperatures. Approximately 90% of farmers relied primarily on pesticides for control of ToLCVD. Few ToLCV-resistant varieties were available but one such variety, Avinash II, was grown by 13% of farmers in the hot season when incidence of the disease is highest. Farmers visiting experimental trials at a farmer field day, showed a strong interest in new advanced breeding tomato lines with resistance to ToLCV. Factors affecting the adoption of ToLCV-resistant varieties by tomato farmers are discussed.     

Animal welfare and ethical issues relevant to the humane control of vertebrate pests

Vertebrate pests and pest control impact on people, animals and the environment, so any ethical consideration of vertebrate pest control must incorporate the interests of all three. The necessity of intervention, whether it involves killing animals or not, must be properly evaluated. Justification for pest control is only tenable if all of the negative impacts (harms) on people, animals and the environment are minimised and all of the positive impacts (benefits) are maximised as far as can be feasibly achieved. In all cases, the most humane control methods possible must be used; we must actively seek ways to improve the humaneness of existing methods and to find new methods that are more humane. There are six major principles that guide the design and execution of ethically sound vertebrate pest control programmes. (1) The aims or benefits and the harms of each control programme must be clear. (2) Control must only be undertaken if the aims can be achieved. (3) The methods that most effectively achieve the aims of the control programme must be used. (4) The methods must be applied in the best possible way. (5) Whether or not each control programme actually achieved its precise aim must be assessed. (6) Once the desired aims or benefits have been achieved, steps must be taken to maintain the beneficial state. An ideal pest control method would be effective and easy to use, affordable, safe for human users and for people exposed to it, humane, specific to the target species or individuals, and safe for the environment. Although such a gold standard is difficult to achieve, we can only retain ethical credibility if we conscientiously strive to make incremental improvements towards that gold standard.     

Canine synovial sarcoma: a retrospective assessment of described prognostic criteria in 16 cases (1994-1999)

Pertinent patient data and biopsied tissue from 16 cases of canine synovial sarcoma (SS) were reviewed. Histopathological grade, clinical stage, and tissue immunoreactivity to cytokeratin (broad stain, AE1/AE3 and cytokeratin 7) and vimentin were determined and correlated with survival. Effect of treatment on survival was similarly evaluated. Neither clinical stage nor histopathological grade significantly affected survival patterns. Tissues from all cases stained >30% positively with vimentin, whereas no tissue from any case exhibited cytokeratin immunoreactivity. Dogs receiving surgical tumor excision or amputation had a significantly higher survivability than those receiving no treatment (P<0.02). Treatment aggressiveness may be more appropriate than clinical staging or tumor grading in predicting survival. Reliability of diagnosing and prognosticating canine SS with current immunohistochemistry protocols should be questioned.     

An unusual presentation of canine distemper virus infection in a domestic ferret (Mustela putorius furo)

A 4.5-year-old, male castrated ferret was examined with a 27-day history of severe pruritus, generalized erythema and scaling. Skin scrapings and a trichogram were negative for mites and dermatophyte organisms. A fungal culture of hair samples was negative. The ferret was treated presumptively for scabies and secondary bacterial and yeast infection with selamectin, enrofloxacin, fluconazole, diphenhydramine and a miconazole–chlorhexidine shampoo. The ferret showed mild improvement in clinical signs over the subsequent 3 weeks, but was inappetent and required supportive feeding and subcutaneous fluids by the owner. The ferret was then examined on an emergency basis at the end of 3 weeks (53 days following initial signs of illness) for severe blood loss from a haematoma over the interscapular region, hypotension and shock. The owners elected euthanasia due to a poor prognosis and deteriorating condition. On post-mortem examination intraepithelial canine distemper viral inclusions were identified systemically, and abundant canine distemper virus antigen was identified with immunohistochemical staining. It is important to note the prolonged course of disease along with the absence of respiratory and neurological signs because this differs from the classic presentation of canine distemper virus infection in ferrets. Canine distemper virus should remain a clinical suspicion for ferrets with skin lesions that do not respond to appropriate therapy, even in animals that were previously vaccinated.     

A retrospective study of canine persistent nasal disease: 80 cases (1998-2003)

Persistent canine nasal disease is a common complaint in small animal practice; however, an etiologic diagnosis can be difficult to establish. The aim of this retrospective study was to determine the percentage of cases for which the etiology was determined in our hospital population. Medical records from 80 dogs met the criteria of inclusion in the study. Nonspecific rhinitis was identified in 23.7% of cases. Other diagnoses were neoplasia (15.0%), fungal infection (nasal aspergillosis) (8.7%), cleft palate (8.7%), periodontal disease (4.0%), parasites (1.3%), foreign body (1.3%), and primary bacterial disease (1.3%). A definitive diagnosis could not be established in 36.3% of cases. Dogs with neoplastic and mycotic diseases often presented with severe radiographic and rhinoscopic lesions. Despite a systematic approach, numerous cases went undiagnosed. The use of advanced imaging should increase our ability to obtain an etiologic diagnosis in canine nasal disease.     

Effects of metaldehyde and acetaldehyde on feeding responses and neuronal activity in the snail,lymnaea stagnalis

The pond snail Lymnaea stagnalis was fed on maize pellets containing metaldehyde. With increasing concentration of metaldehyde there was a decrease in the meal length, number of bites per meal and amount of pellet eaten. The irregularity of biting and the amount of time spent pausing during the meal increased. Meals on maize pellets in the presence of acetaldehyde (one of the metabolites of metaldehyde) were shorter than control meals on maize pellets alone. The symptoms of poisoning after ingestion of metaldehyde or in the presence of acetaldehyde were similar. There was an increase in mucus secretion. Animals showed muscular spasms, undirected mouthing movements and uncoordinated locomotion. This was followed by a period of immobility. The isolated central nervous system of L. stagnalis was used to study the effects of metaldehyde and acetaldehyde on the neural activity underlying feeding. Application of metaldehyde or acetaldehyde led to an increase in firing activity and development of paroxysmal depolarising shifts in buccal motoneurons. It is argued that this type of activity could explain some of the symptoms of poisoning seen in the whole animal after ingestion of metaldehyde, and that acetaldehyde may be responsible for some of the toxic effects of metaldehyde. These results suggest that the CNS of L. stagnalis provides a useful model for screening molluscicides for their effects on a functional neuronal network.     

The fate of the rodenticide flocoumafen in the rat: Retention and elimination of a single oral dose

A single oral dose of 0.14 mg kg^?1 of [¹⁴C] flocoumafen to rat, which gave a transient, non-lethal, effect, was rapidly absorbed, radioactivity appearing in the blood maximally at 4 h and falling to half maximum value by 8 h. The maximum effect on prothrombin time was at 24 h and the value returned to normal by 48 h. Elimination of radioactivity was very slow, with less than 0.5% of the dose in the urine up to 7 days after dosing, and 23-26% in the faeces (more than half of which appeared in the first 24 h). Most of the administered radioactivity (74-76%) was retained 7 days after dosing. Approximately half of the dose was in the liver; it was eliminated with a halflife of 220 days. At 48 h after dosing, most of the hepatic radioactivity comprised unchanged flocoumafen. Treatments of flocoumafen-dosed rats with warfarin or with cytochrome P450-inducing doses of phenobarbitone were without effect on the hepatic residue of flocoumafen.