Hypothesis for heritable, anti-viral immunity in crustaceans and insects

Correction to Flegel, TW: Hypothesis for heritable, anti-viral immunity in crustaceans and insects. Biology Direct 2009, 4:32

Response to Dengue virus infections altered by cytokine-like substances from mosquito cell cultures

<p>Abstract</p> <p>Background</p> <p>With both shrimp and commercial insects such as honey bees, it is known that stable, persistent viral infections characterized by absence of disease can sometimes shift to overt disease states as a result of various stress triggers and that this can result in serious economic losses. The main research interest of our group is to understand the dynamics of stable viral infections in shrimp and how they can be destabilized by stress. Since there are no continuous cell lines for crustaceans, we have used a C6/36 mosquito cell line infected with Dengue virus to test hypotheses regarding these interactions. As a result, we accidentally discovered two new cytokine-like substances in 5 kDa extracts from supernatant solutions of acutely and persistently infected mosquito cells.</p> <p>Results</p> <p>Naïve C6/36 cells were exposed for 48 h to 5 kDa membrane filtrates prepared from the supernatant medium of stable C6/36 mosquito cell cultures persistently-infected with Dengue virus. Subsequent challenge of naïve cells with a virulent stock of Dengue virus 2 (DEN-2) and analysis by confocal immunofluorescence microscopy using anti-DEN-2 antibody revealed a dramatic reduction in the percentage of DEN-2 infected cells when compared to control cells. Similar filtrates prepared from C6/36 cells with acute DEN-2 infections were used to treat stable C6/36 mosquito cell cultures persistently-infected with Dengue virus. Confocal immunofluorescence microscopy revealed destabilization in the form of an apoptosis-like response. Proteinase K treatment removed the cell-altering activities indicating that they were caused by small polypeptides similar to those previously reported from insects.</p> <p>Conclusions</p> <p>This is the first report of cytokine-like substances that can alter the responses of mosquito cells to Dengue virus. This simple model system allows detailed molecular studies on insect cytokine production and on cytokine activity in a standard insect cell line.</p

False rumours of disease outbreaks caused by infectious myonecrosis virus (IMNV) in the whiteleg shrimp in Asia

<p>Abstract</p> <p>Background</p> <p>Infectious myonecrosis virus (IMNV) disease outbreaks in cultivated whiteleg shrimp <it>Penaeus (Litopenaeus) vannamei </it>are characterized by gross signs of whitened abdominal muscles and by slow mortality reaching up to 70%. In 2006 the first disease outbreaks caused by IMNV in Asia occurred in Indonesia. Since then rumours have periodically circulated about IMNV disease outbreaks in other Asian countries. Our findings indicate that these are false rumours.</p> <p>Findings</p> <p>Our continual testing by nested RT-PCR of shrimp samples suspected of IMNV infection from various Asian countries since 2006 has yielded negative results, except for samples from Indonesia. Our results are supported by the lack of official reports of IMNV outbreaks since January 2007 in the Quarterly Report on Aquatic Animal Diseases (QAAD) from the Network of Aquaculture Centers in Asia Pacific (NACA). In most cases, our shrimp samples for which tissue sections were possible showed signs of muscle cramp syndrome that also commonly causes muscle whitening in stressed whiteleg shrimp. Thus, we suspect that most of the false rumours in Asia about IMNV outside of Indonesia have resulted because of muscle cramp syndrome.</p> <p>Conclusions</p> <p>Results from continual testing of suspected IMNV outbreaks in Asian countries other than Indonesia since 2006 and the lack of official country reports of IMNV outbreaks since January 2007, indicate that rumours of IMNV outbreaks in Asian countries outside of Indonesia are false. We suspect that confusion has arisen because muscle cramp syndrome causes similar signs of whitened tail muscles in whiteleg shrimp.</p

A novel integrase-containing element may interact with Laem-Singh virus (LSNV) to cause slow growth in giant tiger shrimp

<p>Abstract</p> <p>Background</p> <p>From 2001-2003 monodon slow growth syndrome (MSGS) caused severe economic losses for Thai shrimp farmers who cultivated the native, giant tiger shrimp, and this led them to adopt exotic stocks of the domesticated whiteleg shrimp as the species of cultivation choice, despite the higher value of giant tiger shrimp. In 2008, newly discovered Laem-Singh virus (LSNV) was proposed as a necessary but insufficient cause of MSGS, and this stimulated the search for the additional component cause(s) of MSGS in the hope that discovery would lead to preventative measures that could revive cultivation of the higher value native shrimp species.</p> <p>Results</p> <p>Using a universal shotgun cloning protocol, a novel RNA, integrase-containing element (ICE) was found in giant tiger shrimp from MSGS ponds (GenBank accession number <ext-link ext-link-id="FJ498866" ext-link-type="gen">FJ498866</ext-link>). <it>In situ </it>hybridization probes and RT-PCR tests revealed that ICE and Laem-Singh virus (LSNV) occurred together in lymphoid organs (LO) of shrimp from MSGS ponds but not in shrimp from normal ponds. Tissue homogenates of shrimp from MSGS ponds yielded a fraction that gave positive RT-PCR reactions for both ICE and LSNV and showed viral-like particles by transmission electron microscopy (TEM). Bioassays of this fraction with juvenile giant tiger shrimp resulted in retarded growth with gross signs of MSGS, and <it>in situ </it>hybridization assays revealed ICE and LSNV together in LO, eyes and gills. Viral-like particles similar to those seen in tissue extracts from natural infections were also seen by TEM.</p> <p>Conclusions</p> <p>ICE and LSNV were found together only in shrimp from MSGS ponds and only in shrimp showing gross signs of MSGS after injection with a preparation containing ICE and LSNV. ICE was never found in the absence of LSNV although LSNV was sometimes found in normal shrimp in the absence of ICE. The results suggest that ICE and LSNV may act together as component causes of MSGS, but this cannot be proven conclusively without single and combined bioassays using purified preparations of both ICE and LSNV. Despite this ambiguity, it is recommended in the interim that ICE be added to the agents such as LSNV already listed for exclusion from domesticated stocks of the black tiger shrimp.</p

Persistent, triple-virus co-infections in mosquito cells

<p>Abstract</p> <p>Background</p> <p>It is known that insects and crustaceans can carry simultaneous, active infections of two or more viruses without showing signs of disease, but it was not clear whether co-infecting viruses occupied the same cells or different cells in common target tissues. Our previous work showed that successive challenge of mosquito cell cultures followed by serial, split-passage resulted in stabilized cultures with 100% of the cells co-infected with Dengue virus (DEN) and an insect parvovirus (densovirus) (DNV). By addition of Japanese encephalitis virus (JE), we tested our hypothesis that stable, persistent, triple-virus co-infections could be obtained by the same process.</p> <p>Results</p> <p>Using immunocytochemistry by confocal microscopy, we found that JE super-challenge of cells dually infected with DEN and DNV resulted in stable cultures without signs of cytopathology, and with 99% of the cells producing antigens of the 3 viruses. Location of antigens for all 3 viruses in the triple co-infections was dominant in the cell nuclei. Except for DNV, this differed from the distribution in cells persistently infected with the individual viruses or co-infected with DNV and DEN. The dependence of viral antigen distribution on single infection or co-infection status suggested that host cells underwent an adaptive process to accommodate 2 or more viruses.</p> <p>Conclusions</p> <p>Individual mosquito cells can accommodate at least 3 viruses simultaneously in an adaptive manner. The phenomenon provides an opportunity for genetic exchange between diverse viruses and it may have important medical and veterinary implications for arboviruses.</p

Emergency response to emerging disease: AHPND in shrimp

Outbreaks of acute hepatopancreatic necrosis disease (AHPND) have caused great economic losses to many shrimp producing countries in Asia since its first appearance in 2009. The causative agent was first reported in 2013 as specific isolates of Vibrio parahaemolyticus (VPAHPND) that were later found to harbor a plasmid (pVA) encoding the Pir-like binary toxin genes PirvpA and PirvpB. More recent information indicates that pVA plasmid and variants occur in many Vibrio parahaemolyticus serotypes and also in other Vibrio species such as V. campbellii, V. harveyi and V. owensii. Information on such genomic and proteomic studies of different VPAHPND isolates from different countries are reviewed. A cohort study carried out in Thailand in 2014 indicated that AHPND outbreaks account for only a portion of the disease outbreaks reported by shrimp farmers as outbreaks of early mortality syndrome (EMS). It is urgent that the etiology of the other EMS-associated mortalities be investigated and not be overlooked. It is recommended that a regional research network and surveillance program for newly-emerging or re-emerging pathogens be established to speed up the process of diagnosis and the implementation of coordinated control measures and to avoid a repeat of the EMS/AHPND scenario

Co-Interactive DNA-Binding between a Novel, Immunophilin-Like Shrimp Protein and VP15 Nucleocapsid Protein of White Spot Syndrome Virus

White spot syndrome virus (WSSV) is one of the most serious pathogens of penaeid shrimp. Although its genome has been completely characterized, the functions of most of its putative proteins are not yet known. It has been suggested that the major nucleocapsid protein VP15 is involved in packaging of the WSSV genome during virion formation. However, little is known in its relationship with shrimp host cells. Using the yeast two-hybrid approach to screen a shrimp lymphoid organ (LO) cDNA library for proteins that might interact with VP15, a protein named PmFKBP46 was identified. It had high sequence similarity to a 46 kDa-immunophilin called FKBP46 from the lepidopteran Spodoptera frugiperda (the fall armyworm). The full length PmFKBP46 consisted of a 1,257-nucleotide open reading frame with a deduced amino acid sequence of 418 residues containing a putative FKBP-PPIase domain in the C-terminal region. Results from a GST pull-down assay and histological co-localization revealed that VP15 physically interacted with PmFKBP46 and that both proteins shared the same subcellular location in the nucleus. An electrophoretic mobility shift assay indicated that PmFKBP46 possessed DNA-binding activity and functionally co-interacted with VP15 in DNA binding. The overall results suggested that host PmFKBP46 might be involved in genome packaging by viral VP15 during virion assembly

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

An overview of PCR techniques for shrimp disease diagnosis in Asia, with emphasis on Thailand

Asia leads the world in cultivated shrimp production with export earnings in the order of billions of US dollars per year. In spite of this success, annual production decreased in the late nineties because of widespread epizootics caused by new viral pathogens. Although, these viruses were no cause for alarm to human health authorities, they were economically crippling for Asian shrimp farmers. In Thailand, shrimp production trends have mirrored those in the rest of Asia, except that recovery from the viral epizootics has been somewhat better than it has for most of its close neighbors. Our work in Thailand has focused on the characterization of the causative viruses and on the development of rapid diagnostic probes for them. Similar work has been done elsewhere. The aim of the work has been to develop effective control measures to help shrimp farmers. We are engaged in similar work on bacteria and parasites. The major viruses of concern (in our estimated order of economic impact for Thailand) are white-spot syndrome virus (WSSV), yellow-head virus (YHV), hepatopancreatic parvovirus (HPV), monodon baculovirus (MBV) and infectious hypodermal and haematopoeitic virus (IHHNV). We have also prepared probes for Vibrio parahaemolyticus and for a microsporidian parasite, Agmasoma penaei. These highly specific and sensitive tools for detection are already helping shrimp farmers and we hope that new technological advances will make them practicable in the field. At the moment, however, the most rapid test is the polymerase chain reaction (PCR) test, which takes approximately 3 hours to complete. This review covers important Asian shrimp diseases for which PCR tests are currently available

Emerging shrimp diseases and innovations to prevent their spread

The number of diseases affecting cultivated penaeid shrimp has increased steadily with expansion and intensification of large-scale commercial cultivation. In the 1990s the most serious losses have occurred from the emergence of new viral pathogens, like white spot virus (WSV) in Asia and Taura syndrome virus (TSV) in the Americas. These initially endemic diseases have quickly become global, probably via infected carriers, including aquaculture stocks. The most alarming of the newly emerging viruses are those like spawner mortality virus (SMV) that produce no pathognomonic lesions and require sophisticated molecular diagnostic techniques for detection. This requirement severely limits the prospects for successful control and it provides a high incentive for the development of more field friendly detection methods. In the coming years, more viral pathogens will undoubtedly be found, especially as cultivation expands geographically and non-native species are used for rearing. To lower risks of disastrous viral epizootics, we should be more cautious with the international movement of living shrimp for aquaculture. We should also lower risks by realistically assessing and limiting other possible modes of pathogen transfer, without unreasonably interfering with international trade. In comparison to viruses, bacteria constitute a much less serious threat to shrimp farmers because most can be controlled by appropriate pond management techniques and because proper chemotherapy is possible in the event of management failure. In spite of this, new pathogens are still being described and too little is known about them. Particularly threatening are new intracellular forms (e.g., molecutes) that may often be overlooked because they cannot be detected in the absence of sophisticated molecular techniques. Even for more traditional pathogens like Vibrio species, we still do not understand why some strains are lethal and others are not. However, some progress is beginning to be made regarding the toxins from Vibrio harveyi and V. penaeicida, the genes controlling toxin production and the possible transfer of these genes amongst strains by bacteriophages. Much remains to be done. To date, fungi and parasites come far behind the viruses and bacteria in the ranking of pathogens that threaten farmed shrimp. However, the example of crayfish plague suggests that this may not always be the case. Control of all these diseases requires a level of understanding that we still lack, not only for the pathogens, but also for the shrimp host. Admirable progress is being made with respect to crayfish cellular defence mechanisms based on pattern recognition proteins targeted against bacterial and fungal cell wall components. Cultivated shrimp species seem to have similar defence systems, but work on them is uneven and much still remains to be confirmed or characterised. In contrast to bacteria, our knowledge of the shrimp response to viral pathogens is almost totally lacking and this is a serious shortcoming given the staggering losses the viruses have caused. We must accelerate investigations at the molecular level for both the host shrimp and its pathogens in order to rationally assess and improve the benefits of proposed therapies and preventative measures for disease. This work must go hand in hand with a concerted effort to develop certified domesticated shrimp stocks and secure, but affordable, cultivation systems