Disease will limit future food supply from the global crustacean fishery and aquaculture sectors.

Seafood is a highly traded food commodity. Farmed and captured crustaceans contribute a significant proportion with annual production exceeding 10 M metric tonnes with first sale value of $40bn. The sector is dominated by farmed tropical marine shrimp, the fastest growing sector of the global aquaculture industry. It is significant in supporting rural livelihoods and alleviating poverty in producing nations within Asia and Latin America while forming an increasing contribution to aquatic food supply in more developed countries. Nations with marine borders often also support important marine fisheries for crustaceans that are regionally traded as live animals and commodity products. A general separation of net producing and net consuming nations for crustacean seafood has created a truly globalised food industry. Projections for increasing global demand for seafood in the face of level or declining fisheries requires continued expansion and intensification of aquaculture while ensuring best utilisation of captured stocks. Furthermore, continued pressure from consuming nations to ensure safe products for human consumption are being augmented by additional legislative requirements for animals (and their products) to be of low disease status. As a consequence, increasing emphasis is being placed on enforcement of regulations and better governance of the sector; currently this is a challenge in light of a fragmented industry and less stringent regulations associated with animal disease within producer nations. Current estimates predict that up to 40% of tropical shrimp production (>$3bn) is lost annually, mainly due to viral pathogens for which standard preventative measures (e.g. such as vaccination) are not feasible. In light of this problem, new approaches are urgently required to enhance yield by improving broodstock and larval sourcing, promoting best management practices by farmer outreach and supporting cutting-edge research that aims to harness the natural abilities of invertebrates to mitigate assault from pathogens (e.g. the use of RNA interference therapeutics). In terms of fisheries losses associated with disease, key issues are centred on mortality and quality degradation in the post-capture phase, largely due to poor grading and handling by fishers and the industry chain. Occurrence of disease in wild crustaceans is also widely reported, with some indications that climatic changes may be increasing susceptibility to important pathogens (e.g. the parasite Hematodinium). However, despite improvements in field and laboratory diagnostics, defining population-level effects of disease in these fisheries remains elusive. Coordination of disease specialists with fisheries scientists will be required to understand current and future impacts of existing and emergent diseases on wild stocks. Overall, the increasing demand for crustacean seafood in light of these issues signals a clear warning for the future sustainability of this global industry. The linking together of global experts in the culture, capture and trading of crustaceans with pathologists, epidemiologists, ecologists, therapeutics specialists and policy makers in the field of food security will allow these issues to be better identified and addressed.

Fungal partnerships stimulate growth of Termitomyces clypeatus stalk mycelium in vitro.

The symbiotic relationship between termites and Termitomyces fungi, which allows the termite to digest cellulose-rich food sources, is poorly understood. In this study, in vitro mixed symbiotic relationships between Termitomyces clypeatus and fungi isolated from individual fungus-comb communities using a culture-dependent method were analyzed. Twenty-day-old stalk cultures of three T. clypeatus isolates were co-cultured with cellulase-producing fungi on potato dextrose agar. The high cellulase-producing fungal isolate no. 18, which showed 99 % ITS sequence identity to Sordariomycetes endophyte isolate 2171 (EU687039), increased growth of T. clypeatus 18/50 by 85.7 %. The high xylanase-producing isolate no. 13, which showed 88 % ITS sequence identity to Arthrinium sacchari isolate L06 (HQ115662), stimulated T. clypeatus 18/50 growth by 58.6 %. The high cellulase- and xylanase-producing isolate no. 50, which showed 90 % ITS sequence identity to the fungal endophyte isolate 2196 (EU687056), improved T. clypeatus 18/50 growth by 45.7 %. A Gigantropanus sp. promoted the growth of T. clypeatus 18/50 and 20/50 by 45.7 and 44.1 %, respectively, and that of T. clypeatus 19/50 by 10.6 %. These results indicated the most beneficial potential partnership of T. clypeatus might involve cellulase-producing fungi isolated from the same ecological niche. The Gigantropanus sp. is a potential partner of T. clypeatus but is likely to be less common than cellulase-producing fungi isolated from fungus combs owing to the lower host specificity of the Gigantropanus sp. This study provides an interesting method to culture Termitomyces using an in vitro mixed culture method for production of Termitomyces fruiting bodies in the future.

Clinical use of a new frameless optical neuronavigation system for brain biopsies: 10 cases (2013-2020).

OBJECTIVES: The aim of the retrospective study was to describe the brain biopsy procedure using a new frameless optical neuronavigation system and to report diagnostic yield and complications associated with the procedure. MATERIALS AND METHODS: The medical records for all dogs with forebrain lesions that underwent brain biopsy with a frameless optical neuronavigation system in a single referral hospital between 2013 and 2020 were retrospectively analysed. Following data were collected: signalment, neurological signs, diagnostic findings, number of brain biopsy samples, sampled region, complications, duration of hospitalisation, whether the samples were diagnostic and histopathological diagnoses. The device consists of a computer workstation with navigation software, an infrared camera, patient tracker and reflective instruments. The biopsy needle was equipped with reflective spheres, so the surgeon could see the position of the needle during sampling the intracranial lesion free handed through a mini-burr hole. RESULTS: Ten dogs were included. Absolute diagnostic yield based on specific histopathological diagnosis was 73.9%. Three dogs had immune-mediated necrotizing encephalitis, two dogs showed a necrotizing leukoencephalitis and two dogs a meningoencephalitis of unknown origin. In two dogs, the brain specimen showed unspecific changes. In one dog, the samples were non-diagnostic. Seven dogs showed no neurological deterioration, one dog mild temporary ataxia and two dogs died within 36 hours post brain biopsy. CLINICAL SIGNIFICANCE: In these 10 dogs, the frameless optical neuronavigation system employed was useful to gain diagnostic brain biopsy samples. Considering the mortality rate observed, further studies are needed to confirm the safety of this procedure and prove its actual clinical effectiveness.

Identification and characterization of Alix/AIP1 interacting proteins from the black tiger shrimp, Penaeus monodon.

Apoptosis is proposed to be a major cause of death in shrimp viral infections. From our previous study, an apoptosis-related gene, Pm-Alix, was identified from the black tiger shrimp. Its expression was high in defence-related tissues including haemocytes and the lymphoid organ. To clarify its possible role in shrimp, we used Pm-Alix as bait in a yeast two-hybrid analysis to search for Alix interacting proteins in shrimp. Two cDNA sequences discovered had homology to a predicted ubiquitin C of the purple sea urchin, Strongylocentrotus purpuratus, and to a guanylyl cyclase of the red swamp crayfish, Procambarus clarkii. In vitro pull-down assays confirmed positive interaction between Pm-Alix and both proteins. Tissue distribution analysis revealed that Pm-Alix and the two binding partners were widely expressed in various tissues but more highly expressed in haemocytes. However, no significant positive or negative correlation was found in the expression of these genes as shrimp approached morbidity and death after challenge with white spot syndrome virus. Thus, the results suggested that Alix and its interacting partners did not play a direct role related to shrimp death.

Impact of yellow head virus outbreaks in the whiteleg shrimp, Penaeus vannamei (Boone), in Thailand.

Yellow head virus (YHV) is known as a major pathogen in the black tiger shrimp, Penaeus (Penaeus) monodon. It can also cause serious mortality in farmed whiteleg shrimp, Penaeus (Litopenaeus) vannamei. However, there is no published information on the economic and/or production impact of the disease in P. vannamei. Shrimp with gross signs of YHV disease (faded body colour and 60-70% mortality) were observed in 20 study farms rearing P. vannamei in the central part of Thailand from the end of 2007 through early 2008. The estimated economic loss for these farms according to the Thai Animal Aquaculture Association was approximately US$3 million. Detailed sequence analysis of RT-PCR amplicons from shrimp in all the study ponds revealed the presence of YHV Type 1b (YHV-1b) alone (characterized by a 162-bp deletion in the ORF3 region encoding the structural gene for gp116) and the absence of YHV Type 1a (YHV-1a), the original YHV type reported from Thailand. Despite the large 162-bp deletion (= 54 deduced amino acids) in the gp116 structural gene, histopathology of YHV-1b infections was identical to that of YHV-1a infections, and electron microscopy revealed that YHV-1b virions were morphologically indistinguishable from those previously reported for YHV-1a. In addition, an existing commercial RT-PCR detection kit and an immunochromatographic test strip for the detection of YHV were proven to have been valid tests for both YHV-1b and YHV-1a. The source of the virus for these outbreaks was unlikely to have been the post-larvae used to stock the ponds, as they were derived from domesticated specific pathogen-free stocks free of YHV. Thus, it is possible that they originated from an unknown, natural reservoir.

Research progress on viral accommodation 2009 to 2019.

The viral accommodation hypothesis for crustaceans and insects was first proposed in 1998/2001, stimulated by observations that shrimp and insects or insect cell lines can coexist with both DNA or RNA viruses without showing any signs of disease (i.e., they tolerate, single to multiple, persistent infections, sometimes for a lifetime). A review of tests of the hypothesis up to 2007 was previously published in DCI. This was followed by a major revision in 2009 when the elusive memory element required by the hypothesis was proposed to reside in non-retroviral fragments of extant viruses, now called endogenous viral elements (EVE) that are autonomously inserted into the host genome as cDNA copied from viral mRNA. Here, progress in research on viral accommodation in crustaceans and insects over the decade following 2009 is reviewed. It culminates with a discussion of exiting research results from insects in 2019 that prove the existence of specific, adaptive and heritable immunity, at least in mosquitoes. It remains to be determined whether the same mechanisms also govern EVE acquisition and its protective RNA production in shrimp. The wide-ranging consequences of the revealed mechanisms for viral disease control in economic crustaceans and insects is discussed.

Magnetic resonance imaging characteristics of necrotizing meningoencephalitis in Pug dogs.

BACKGROUND: The magnetic resonance imaging (MRI) characteristics of necrotizing meningoencephalitis (NME) are not well documented. OBJECTIVES: To describe common MRI features of NME, to compare the MRI features to histopathologic findings, and to determine whether or not MRI lesions are predictive of survival time. ANIMALS: Eighteen Pugs with NME. METHODS: Retrospective MRI case study of Pugs identified by a search of medical records at 6 veterinary institutions. Eighteen dogs met inclusion criteria of histopathologically confirmed NME and antemortem MRI exam. MRI lesions were characterized and compared with histopathology with the kappa statistic. Survival times were compared with MRI findings by use of Mann-Whitney U-tests and Spearman's rho. RESULTS: Twelve of 18 lesions were indistinctly marginated with mild parenchymal contrast enhancement. Prosencephalic (17/18) lesion distribution included the parietal (16/18), temporal (16/18), and occipital (16/18) lobes. There were cerebellar (4/18) and brainstem (3/18) lesions. Asymmetric lesions were present in both gray and white matter in all dogs. Falx cerebri shift was common (11/18), and 6 dogs had brain herniation. Leptomeningeal enhancement was present in 9/18 dogs. A moderate positive association was found between parenchymal contrast enhancement and both necrosis (kappa= 0.45; P= .045) and monocytic inflammation (kappa= 0.48; P= .025). Higher MRI lesion burden was correlated with longer time from disease onset to MRI (P= .045). MRI lesion burden did not correlate to survival time. CONCLUSIONS AND CLINICAL IMPORTANCE: Asymmetric prosencephalic grey and white matter lesions with variable contrast enhancement were consistent MRI changes in Pugs with confirmed NME. While not pathognomonic for NME, these MRI characteristics should increase confidence in a presumptive diagnosis of NME in young Pugs with acute signs of neurologic disease.

Shrimp hepatopancreatic parvovirus detection by combining loop-mediated isothermal amplification with a lateral flow dipstick.

Present methods such as traditional PCR, PCR-ELISA, real-time PCR and histopathology for detection of shrimp hepatopancreatic parvovirus (PmDNV) entail various disadvantages including high cost, long assay time or use of toxic substances. Loop-mediated isothermal amplification (LAMP) of target nucleotide sequences under inexpensive isothermal conditions combined with amplicon detection by chromatographic lateral flow dipsticks (LFD) allowed simpler detection within 75 min. Biotinylated LAMP amplicons from the targeted portion of the PmDNA capsid protein gene were produced under isothermal conditions at 63 degrees C for 1h and then hybridized at 63 degrees C for 5 min with an FITC-labeled probe (optimized at 20 pmol) that was specific for the LAMP amplicons (i.e., outside the primer region). The FITC-labeled, biotinylated LAMP product picked up gold-labeled, anti-FITC near the LFD origin and the whole, triple-labeled complex was captured by an immobilized biotin-binding protein to yield a red nano-gold stripe at the LFD test line. With a DNA template derived from PmDNV-infected shrimp, the LAMP-LFD detection limit was 1 ng while that for one-step PCR-electrophoresis was 10 ng. Comparative sensitivity for one nested-PCR-electrophoresis method was 1 ng but for another 0.1 ng. The LAMP-LFD method gave negative test results with DNA extracts from normal shrimp and from shrimp infected with other DNA viruses including monodon baculovirus (MBV), white spot syndrome virus (WSSV) and infectious hypodermal and hematopoietic necrosis virus (IHHNV).

Shrimp Taura syndrome virus detection by reverse transcription loop-mediated isothermal amplification combined with a lateral flow dipstick.

Loop-mediated isothermal amplification (LAMP) allows rapid amplification of nucleic acid under isothermal conditions using four sets of specially designed primers that recognize six distinct target sequences with high specificity and sensitivity. In this report, a 60-min reverse transcription LAMP (RT-LAMP) method for amplification of Taura syndrome virus (TSV) cDNA using biotin-labeled primer was combined with a chromatographic lateral flow dipstick (LFD) for rapid and simple visual detection of TSV-specific amplicons. The LFD process involved a 5-min post RT-LAMP step for specific hybridization of cDNA with an FITC-labeled DNA probe that confirmed the presence of specific, biotin-labeled TSV amplicons. The resulting DNA duplexes could be visualized trapped at the LFD strip test line within 5min of sample exposure. Using the combined RT-LAMP and LFD system, the total assay interval was approximately 70min, excluding RNA extraction time. Detection sensitivity was comparable to other commonly used methods for nested RT-PCR detection of TSV. In addition to reduced assay time when compared to electrophoresis, combination of RT-LAMP with LFD confirms amplicon identity by hybridization and eliminates the need to handle carcinogenic ethidium bromide.

Computer-assisted magnetic resonance imaging brain morphometry in American Staffordshire Terriers with cerebellar cortical degeneration.

BACKGROUND: Cerebellar cortical degeneration exists in American Staffordshire Terriers. Magnetic resonance imaging (MRI) can be suggestive, but a definitive diagnosis requires histopathology. HYPOTHESIS: Computer-assisted MRI morphometry can be used to distinguish between American Staffordshire Terriers with or without cerebellar cortical degeneration. ANIMALS: Normal American Staffordshire Terriers (n = 17) and those with clinical signs of cerebellar cortical degeneration (n = 14). METHODS: This was a partly retrospective and partly prospective study. Causes of cerebellar disease were ruled out with brain MRI, cerebrospinal fluid (CSF) analysis, CBC, blood biochemistry, and clinical follow-up. On T2-weighted midsagittal MR images, the following parameters were calculated: size of the cerebellum relative to the entire brain, size of the CSF space surrounding the cerebellum relative to the cerebellum, and 2 threshold-dependent cerebellar CSF indices (with and without surrounding CSF). RESULTS: Statistical analyses indicated a significantly lower relative cerebellar size (P < .001) and a larger relative cerebellar CSF space (P < .001) in dogs with cerebellar cortical degeneration. The measurement of relative cerebellar size could distinguish between affected and nonaffected dogs with a sensitivity and a specificity of 93 and 94%, respectively, using a cut-off of 13.3%. Using a cut-off of 12.8%, the measurement of relative CSF space could distinguish between both groups with a sensitivity of 93% and a specificity of 100%. There was a significant difference in 1 of the 2 CSF indices between affected and normal dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: Relative cerebellar size and relative CSF space calculated from MRI are effective in American Staffordshire Terriers to differentiate between normal animals and those with cerebellar cortical degeneration.

Sequential magnetic resonance imaging changes after suspected global brain ischaemia in a dog.

Sequential magnetic resonance imaging studies over a period of 18 months were performed in a two-year-old pug dog after suspected global brain ischaemia following an anaesthetic accident. The dog was presented with seizures and neurological deficits consistent with a left brainstem and multifocal/diffuse forebrain lesion after an asymptomatic interval of 72 hours following the ischaemic event. Magnetic resonance imaging scans were performed three hours, six weeks, seven and 18 months after the incident. In the acute stage, signal hyperintensity was evident in the occipital and parietal regions of the cerebral cortex and in both rostral caudate nuclei. Slight involvement of the white matter was also noticed. In the chronic phase, the signal hyperintensity in the affected areas of the cortex was diminished and smaller in size, whereas the white matter did not appear to be compromised anymore.

Canine NAPEPLD-associated models of human myelin disorders.

Canine leukoencephalomyelopathy (LEMP) is a juvenile-onset neurodegenerative disorder of the CNS white matter currently described in Rottweiler and Leonberger dogs. Genome-wide association study (GWAS) allowed us to map LEMP in a Leonberger cohort to dog chromosome 18. Subsequent whole genome re-sequencing of a Leonberger case enabled the identification of a single private homozygous non-synonymous missense variant located in the highly conserved metallo-beta-lactamase domain of the N-acyl phosphatidylethanolamine phospholipase D (NAPEPLD) gene, encoding an enzyme of the endocannabinoid system. We then sequenced this gene in LEMP-affected Rottweilers and identified a different frameshift variant, which is predicted to replace the C-terminal metallo-beta-lactamase domain of the wild type protein. Haplotype analysis of SNP array genotypes revealed that the frameshift variant was present in diverse haplotypes in Rottweilers, and also in Great Danes, indicating an old origin of this second NAPEPLD variant. The identification of different NAPEPLD variants in dog breeds affected by leukoencephalopathies with heterogeneous pathological features, implicates the NAPEPLD enzyme as important in myelin homeostasis, and suggests a novel candidate gene for myelination disorders in people.

Update on viral accommodation, a model for host-viral interaction in shrimp and other arthropods.

Comparatively little published information is available on the mechanistic response of shrimp and other arthropods to viral pathogens. Much of the literature has been focused on the use of viruses for biological control of insect pests or disease vectors and on the use of baculoviruses as a means of heterologous protein expression in insect cell lines. The situation changed dramatically with the rapid global increase in cultivation of penaeid shrimp and the massive farm losses that have occurred due to viral pathogens. Urgency to solve these problems has led to a closer examination of the shrimp response to viral pathogens in the hope of finding new methods of disease control. Field observations and results of laboratory experiments in the past decade indicate that shrimp may be capable of a specific, adaptive response to viral pathogens that cannot be explained by current knowledge and understanding of their cellular and humoral defenses. Hallmarks of this response are specific tolerance to single and multiple viral infections without gross or histological signs of disease, a phenomenon common to crustaceans and insects. The concept of viral accommodation was introduced in 1998 as a simple testable hypothesis to explain these phenomena. Key elements of the hypothesis were an unknown mechanism for specific memory of pathogens and the role of this memory in dampening viral triggered apoptosis. Recent field and research results have supported predictions of the viral accommodation hypothesis and suggest that memory may be provided by the viral pathogens themselves in persistent infections that result in reduced severity of disease. The well-known phenomenon of defective interfering viral particles may play an important role in this process, but it cannot explain cross protection that has recently been described for heterologous viral infections. The major conclusion is that homologous and heterologous reduction in disease severity resulting from persistent viral infections (i.e., accommodated viral infections) may be a key process that has evolved from host viral interaction in the arthropod line.

Rapid and sensitive detection of Taura syndrome virus by reverse transcription loop-mediated isothermal amplification.

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay is a novel method of gene amplification that amplifies nucleic acid with high specificity, sensitivity and rapidity, which can be applied for disease diagnosis in shrimp aquaculture. The method is performed under isothermal conditions with a set of four specially designed primers that recognize six distinct sequences of the target. In this study, using the RT-LAMP method, a protocol for detecting Taura syndrome virus which is a causative agent of Penaeus vannamei was developed. Time and temperature conditions for detection of TSV were optimized for 60min at 63 degrees C. The nucleic acids of other shrimp pathogens (yellow head virus; YHV and white spot syndrome; WSSV) were not amplified by this RT-LAMP system. The detection of TSV using RT-LAMP was 10 times more sensitive than the RT-PCR but less sensitive than nested RT-PCR. However this system was more convenient, rapid, and does not require sophisticated PCR machine.

Cerebellar cortical degeneration with selective granule cell loss in Bavarian mountain dogs.

Three Bavarian mountain dogs aged between 18 and 20 months, not related to each other, were presented with chronic signs of cerebellar dysfunction. On sagittal T2-weighted magnetic resonance imaging brain images, the tentative diagnosis of cerebellar hypoplasia was established based on an enlarged cerebrospinal fluid space around the cerebellum and an increased cerebrospinal fluid signal between the folia. Post-mortem examination was performed in one dog and did show an overall reduction of cerebellar size. On histopathologic examination, a selective loss of cerebellar granule cells with sparing of Purkinje cells was evident. Therefore, the Bavarian mountain dog is a breed where cerebellar cortical degeneration caused by the rather exceptional selective granule cell loss can be seen as cause of chronic, slowly progressive cerebellar dysfunction starting at an age of several months.

Relapses in dogs with steroid-responsive meningitis-arteritis.

OBJECTIVES: To describe relapse rates in steroid-responsive meningitis-arteritis and to describe clinical and laboratory parameters in dogs with and without relapses. METHODS: Seventy-four dogs with steroid-responsive meningitis-arteritis were retrospectively identified and assigned to one of three groups: (1) without relapse; (2) at least one relapse and (3) unknown relapse status. The following parameters are reported for the first two groups: sex, age, breed, body weight, nucleated cell count, total protein concentration and percentage of neutrophils on initial cerebrospinal fluid analysis, immunoglobulin A in serum and initial cerebrospinal fluid analysis, nucleated cell count on cerebrospinal fluid analysis at 3-month re-evaluation, C-reactive protein in serum and initial cerebrospinal fluid analysis and at 3-month re-evaluation. RESULTS: Relapses occurred in 32 · 4% of dogs (one relapse: 62 · 5%; two relapses: 25 · 0%; three relapses: 8 · 3%; four relapses: 4 · 2%), 55 · 4% were relapse-free and in 12 · 2% the relapse status was unknown. C-reactive protein in serum and cerebrospinal fluid on 3-month re-evaluation was normal in 80% and 75% of dogs with relapses, respectively. In dogs without relapse, C-reactive protein in serum and cerebrospinal fluid on 3-month re-evaluation was normal in 100% and 90% of dogs, respectively. CLINICAL SIGNIFICANCE: Relapses are frequent but no reliable predictive indicator has emerged in this study. Nevertheless, elevated C-reactive protein in serum warrants continuing therapy; normal C-reactive protein in serum does not exclude future relapse.

Target for standard Thai PCR assay identical in 12 white spot syndrome virus (WSSV) types that differ in DNA multiple repeat length.

A Thai PCR detection method (WSSV-232) yielding a 232 bp amplicon has been used for detection of white spot syndrome virus (WSSV) since 1996. It targets ORF 91 in the full sequence of the only Thai WSSV isolate at GenBank (AF369029). At the beginning of 2002, some Thai shrimp farmers complained that ponds stocked with WSSV-232 PCR negative post-larvae (PL) later suffered WSSV disease outbreaks. Although these outbreaks may have resulted from horizontal transmission of WSSV after stocking, it was also possible that they resulted from false negative PCR test results due to genetic changes at the PCR-assay target after the first appearance of WSSV in Thailand in 1995. Indeed, recent results have revealed at least 12 WSSV variants in Thailand that can be distinguished based on differences in DNA multiple repeat lengths in ORF 94 (GenBank AF369029). To test for variation in the WSSV-232 target sequence in ORF 91, 20 DNA extracts derived from field samples and representing 9 of the WSSV DNA multiple repeat groups were subjected to PCR amplification and sequencing using primers that generated a 403 bp amplicon covering the target for the WSSV-232 assay. An additional three repeat types were included from archived material. Analysis revealed that the 232 bp target sequence in ORF 91 was unchanged in all of the 12 types tested and that the original WSSV-232 detection system was still valid. Thus, any false negative PCR test results leading to farmer complaints would probably have arisen from small sample sizes and low sensitivity of the single-step PCR assay. If so, false negative results could be reduced by the use of nested PCR assays with larger PL sample sizes. .

Comparison of penaeid shrimp and insect parvoviruses suggests that viral transfers may occur between two distantly related arthropod groups.

The DNA and putative amino acid sequences of representative insect and shrimp parvoviruses (subfamily Densovirinae) were analyzed using computer programs. Shrimp viruses included hepatopancreatic parvovirus (HPV) of Penaeus monodon (HPVmon) and P. chinensis (HPVchin), spawner-isolated mortality virus from P. monodon (SMVmon) and infectious hypodermal and hematopoietic necrosis virus (IHHNV) from P. vannamei. Insect viruses included Aedes aegypti densovirus (AaeDNV), Aedes albopictus densovirus (AalDNV), Junonia coenia densovirus (JcDNV), Galleria mellonella densovirus (GmDNV), Bombyx mori densovirus 5 (BmDNV), Diatraea saccharalis densovirus (DsDNV) and Periplaneta fuliginosa densovirus (PfDNV). Virion size for all these viruses ranged between 18 and 30 nm diameter and ssDNA genome length was between 4 and 6 kb. Using BLAST or Clustal W with the sequence fragments available, no significant DNA homology was found except for 77% DNA identity between HPVmon and HPVchin. However, phylogenetic trees constructed by comparing DNA genome sequences for putative viral polypeptides, capsid proteins and nonstructural proteins placed the parvoviruses into two Clades: Clade 1 with SMVmon, PfDNV, DsDNV, GmDNV, JcDNV, and BmDNV; and Clade 2 with HPVmon, HPVchin, IHHNV, AalDNV and AaeDNV. The four shrimp parvoviruses fell into two different clades that grouped with different insect parvoviruses.

Structure elucidation of glykenin glycosidic antibiotics from Basidiomycetes sp. V. High-performance liquid chromatographic separation of components of glykenin.

The glycosidic antibiotics of the glykenin (GK) family produced by Basidiomycetes sp. were separated into nine components (GK-I-VII and DG) by normal-phase chromatography. It was found that these components differ in the number and location of the acetyl groups in the sugar moiety. Each component (GK-I-VII and DG) was further separated into three isomers (A, B and C), which possess different aglycones, by reversed-phase chromatography on an ODS column with methanol-acetonitrile as eluent. The best composition of the eluent was found to be methanol-acetonitrile-1% trifluoroacetic acid (4:3.5:2.5). The profile analysis of GK-III-VII and DG was also carried out using a modified mobile phase. The combination of normal- and reversed-phase chromatography separated all components of the GK mixture except GK-I and II. The relationship between structure and separation behaviour of GK is discussed.