Silver nanoparticles induce histopathological alterations in juvenile Penaeus vannamei.

The objective of this study was to evaluate the histopathological alterations in juvenile Penaeus vannamei caused by silver nanoparticles (AgNPs) for two types of experiments: at sublethal concentrations of 3.6 to 7.1 µg/µL of metallic silver (Ag) for a short period up to 72 h and for 2.6 to 7.9 µg of Ag/µL for the long period up to 264 h. The severity degree of the changes was evaluated and the histopathological index (Hi) was determined in both experiments using the necrosis (cellular dead) as an indicator. The pathological changes in the striated muscle, gills, antennal gland, circulatory system, heart, lymphoid organ, and connective tissue are described. The histopathological effects were similar for the two experiments without a direct relationship with the concentrations. In the short-term experiment, the values of Hi were higher (2.34 ± 0.41 at 48 hpi and 1.91 ± 0.39 at 72 hpi) compared with the long-term experiment (values between 0.57 ± 0.36 to 1.74 ± 0.57 at 264 hpi). The observed pathologies are similar to those caused by other metals, with the exception of the agglomerations of black particles in the gills, lymphoid organ, and muscle, which has not been previously reported. This work shows that silver nanoparticles cause damage to shrimp in sublethal concentrations.

Silver nanoparticles enhance survival of white spot syndrome virus infected Penaeus vannamei shrimps by activation of its immunological system.

The global aquaculture has shown an impressive growth in the last decades contributing with a major part of total food fish supply. However, it also helps in the spread of diseases that in turn, causes great economic losses. The White Spot Syndrome Virus (WSSV) is one of the major viral pathogen for the shrimp aquaculture industry. Several attempts to eliminate the virus in the shrimp have been addressed without achieving a long-term effectiveness. In this work, we determine the capacity of the commercial non-toxic PVP-coated silver nanoparticles to promote the response of the immune system of WSSV-infected shrimps with or without an excess of iron ions. Our results showed that a single dose of metallic silver in the nanomolar range (111 nmol/shrimp), which is equivalent to 12 ng/mL of silver nanoparticles, produces 20% survival of treated infected shrimps. The same concentration administered in healthy shrimps do not show histological evidence of damage. The observed survival rate could be associated with the increase of almost 2-fold of LGBP expression levels compared with non-treated infected shrimps. LGBP is a key gene of shrimp immunological response and its up-regulation is most probably induced by the recognition of silver nanoparticles coating by specific pathogen-associated molecular pattern recognition proteins (PAMPs) of shrimp. Increased LGBP expression levels was observed even with a 10-fold lower dose of silver nanoparticles (1.2 ng/shrimp, 0.011 nmol of metallic silver/shrimp). The increase in LGBP expression levels was also observed even in the presence of iron ion excess, a condition that favors virus proliferation. Those results showed that a single dose of a slight amount of silver nanoparticles were capable to enhance the response of shrimp immune system without toxic effects in healthy shrimps. This response could be enhanced by administration of other doses and might represent an important alternative for the treatment of a disease that has still no cure, white spot syndrome virus.

Effect of silver nanoparticles on the metabolic rate, hematological response, and survival of juvenile white shrimp Litopenaeus vannamei.

White spot syndrome virus (WSSV) is highly lethal and contagious in shrimps; its outbreaks causes an economic crisis for aquaculture. Several attempts have been made to treat this disease; however, to date, there is no effective cure. Because of their antimicrobial activities, silver nanoparticles (AgNPs) are the most studied nanomaterial. Although the antiviral properties of AgNPs have been studied, their antiviral effect against viral infection in aquaculture has not been reported. The AgNPs tested herein are coated with polyvinylpyrrolidone (PVP) and possess multiple international certifications for their use in veterinary and human applications. The aim of this work was to evaluate the survival rate of juvenile white shrimps (Litopenaeus vannamei) after the intramuscular administration of AgNPs. For this, different concentrations of metallic AgNPs and PVP alone were injected into the organisms. After 96 h of administration, shrimp survival was more than 90% for all treatments. The oxygen consumption routine rate and total hemocyte count remained unaltered after AgNP injection, reflecting no stress caused. We evaluated whether AgNPs had an antiviral effect in shrimps infected with WSSV. The results revealed that the survival rate of WSSV-infected shrimps after AgNP administration was 80%, whereas the survival rate of untreated organisms was only 10% 96 h after infection. These results open up the possibility to explore the potential use of AgNPs as antiviral agents for the treatment of diseases in aquaculture organisms, particularly the WSSV in shrimp culture.

Double-stranded RNA against white spot syndrome virus (WSSV) vp28 or vp26 reduced susceptibility of Litopenaeus vannamei to WSSV, and survivors exhibited decreased susceptibility in subsequent re-infections.

The antiviral effect of vp28 or vp26 double-stranded (ds) RNA upon single or consecutive white spot syndrome virus (WSSV) intramuscular challenges with a high infectious dose was evaluated. The vp28 dsRNA showed the highest protection both in single (LT(50)=145h at 10d and 98h at 20d post treatment [dpt]) or consecutive (LT(50)=765h) WSSV challenges compared to vp26 dsRNA (LT(50)=126h at 10 d and 57h at 20dpt vs. consecutive challenge LT(50)=751h). Single WSSV challenges showed that animals treated with vp28 or vp26 dsRNA gradually lost the antiviral effect as virus challenge occurred at 10dpt (cumulative mortality 63% vs. 80%, respectively) or 20dpt (87% vs. 100%, respectively). In contrast, animals treated with vp28 or vp26 dsRNA and consecutively challenged with WSSV showed and extended lower susceptibility to WSSV. All dead animals were WSSV-positive by one-step PCR, whereas all surviving shrimp from single or continuous challenges were WSSV-negative as determined by reverse transcription (RT)-PCR. In conclusion, shrimp treated with a single administration of vp28 or vp26 dsRNA and consecutively challenged with WSSV showed a stronger and longer antiviral response than shrimp exposed once to WSSV at 10 or 20dpt.