Ovine fetal immune response to Cache Valley virus infection.

Cache Valley virus (CVV)-induced malformations have been previously reproduced in ovine fetuses. To evaluate the development of the antiviral response by the early, infected fetus, before the development of immunocompetency, ovine fetuses at 35 days of gestation were inoculated in utero with CVV and euthanized at 7, 10, 14, 21, and 28 days postinfection. The antiviral immune response in immature fetuses infected with CVV was evaluated. Gene expression associated with an innate, immune response was quantified by real-time quantitative PCR. The upregulated genes in infected fetuses included ISG15, Mx1, Mx2, IL-1, IL-6, TNF-α, TLR-7, and TLR-8. The amount of Mx1 protein, an interferon-stimulated GTPase capable of restricting growth of bunyaviruses, was elevated in the allantoic and amniotic fluid in infected fetuses. ISG15 protein expression was significantly increased in target tissues of infected animals. B lymphocytes and immunoglobulin-positive cells were detected in lymphoid tissues and in the meninges of infected animals. These results demonstrated that the infected ovine fetus is able to initiate an innate and adaptive immune response much earlier than previously known, which presumably contributes to viral clearance in infected animals.

Identification of the target cells and sequence of infection during experimental infection of ovine fetuses with Cache Valley virus.

Cache Valley virus-induced malformations have been previously reproduced in ovine fetuses; however, no studies have established the course of infection of cells and tissues with Cache Valley virus. To address these questions, ovine fetuses at 35 days of gestation were inoculated in utero with Cache Valley virus and euthanized at 7, 10, 14, 21, and 28 days postinfection. On postmortem examination, arthrogryposis and oligohydramnios were observed in some infected fetuses. Morphological studies showed necrosis in the central nervous system and skeletal muscle of infected fetuses evaluated after 7 to 14 days postinfection, and hydrocephalus, micromyelia, and muscular loss were observed in infected fetuses after 21 to 28 days postinfection. Using immunohistochemistry and in situ hybridization, intense Cache Valley virus antigen and RNA staining was detected in the brain, spinal cord, skeletal muscle, and, to a lesser degree, in fetal membranes and other tissues of infected fetuses. Viral antigen and RNA staining decreased in targeted and infected tissues with the progression of the infection.

Streptococcosis in farmed Litopenaeus vannamei: a new emerging bacterial disease of penaeid shrimp.

Presumptive systemic streptococcal infections were detected histologically in farmed Litopenaeus vannamei juveniles submitted from a Latin American country and the bacteria isolated. Characterization work demonstrated that the Gram-positive cocci form chains, grow aerobically and anaerobically, are oxidase- and catalase-negative, non-hemolytic, non-motile, Lancefield Group B positive and PCR positive when amplified with a universal streptococcal primer set. Differing Streptococcus identifications were obtained using API 20 Strep and Biolog systems, the former identifying the isolate as S. uberis and the latter as S. parauberis. Injection of specific pathogen-free (SPF) L. vannamei with the bacteria resulted in 100% mortality by 3 d post-injection with successful recovery of the agent from moribund test shrimp hemolymph samples. The recovered isolate was used in per os and waterborne exposure studies of SPF L. vannamei with mortalities ranging from 40 to 100% and 80 to 100%, respectively. Histologic analysis of 5 to 8 moribund shrimp from each exposure method demonstrated that all contained a severe bacteremia characterized by numerous free cocci within the hemolymph and aggregates of vacuolated hemocytes with notable intravacuolar cocci. This unique lesion type was most pronounced within the lymphoid organ and considered pathodiagnostic for this disease. Experimentally induced lesions were identical to those in naturally infected farmed shrimp and the Streptococcus sp. responsible was re-isolated, fulfilling Koch's postulates. Five freeze/thaw cycles of 10 experimentally infected shrimp were performed over a 2 mo period and the bacteria successfully cultured from all shrimp at each interval. These collective findings describe the first reported case of streptococcosis in marine penaeid shrimp in the Western Hemisphere and indicate that the agent may be disseminated via live or frozen infected shrimp.