Sweat Production During Continuous and Interval Aerobic Exercise.

INTRODUCTION:Aerobic exercise within the habitable volume of small spacecraft needed for space exploration beyond low Earth orbit is expected to challenge the capacity of environmental control systems. Moisture control is a primary concern. Crewmembers will contribute moisture to the cabin environment in the form of sweat while exercising. The effects of continuous aerobic exercise for improving and maintaining aerobic capacity is well characterized. Likewise, evidence suggests that high intensity interval exercise for shorter durations is also effective in building and maintaining aerobic capacity.METHODS: On separate days, measures of sweat and respiratory responses were made for continuous (30 min of steady state exercise at ∼75% of aerobic capacity) and two interval (4 × 4 min, 8 × 30 s) exercise protocols.RESULTS: We observed that the 4-min and 30-s interval protocols produce 16% and 66% less metabolic water loss vs. the continuous exercise protocol, respectively. These responses were highly correlated with the amount of work performed (R² = 0.81) and the amount of energy expenditure (R² = 0.83) during exercise.DISCUSSION: These results suggest that interval exercise may be a useful alternative to continuous aerobic exercise when metabolic water production is an environmental concern. The results may inform the choices of aerobic exercise countermeasure protocols for use in deep space exploration.Ryder JW, Crowell JB, Song HJ, Ewert M. Sweat production during continuous and interval aerobic exercise. Aerosp Med Hum Perform. 2023; 94(8):623-628.

Genetic diversity of porcine reproductive and respiratory syndrome virus in Korea.

The high genetic diversity of porcine reproductive and respiratory syndrome virus (PRRSV) has been an obstacle to developing an effective vaccine for porcine reproductive and respiratory syndrome (PRRS). This study was performed to assess the degree of genetic diversity among PRRSVs from Korean pig farms where wasting and respiratory syndrome was observed from 2005 to 2009. Samples from 786 farms were tested for the presence of PRRSV using reverse transcription PCR protocol. A total of 117 farms were positive for type 1 PRRSV while 198 farms were positive for type 2. Nucleotide sequences encoding the open reading frame (ORF) 5 were analyzed and compared to those of various published PRRSV isolates obtained worldwide. Sequence identity of the ORF 5 in the isolates was 81.6˜100% for type 1 viruses and 81.4˜100% for type 2 viruses. Phylogenetic analysis of the ORF 5 sequences showed that types 1 and 2 PRRSVs from Korea were mainly classified into three and four clusters, respectively. The analyzed isolates were distributed throughout the clusters independent of the isolation year or geographical origin. In conclusion, our results indicated that the genetic diversity of PRRSVs from Korean pig farms is high and has been increasing over time.

Protective immunity induced by systemic and mucosal delivery of DNA vaccine expressing glycoprotein B of pseudorabies virus.

A murine model immunized by systemic and mucosal delivery of plasmid DNA vaccine expressing glycoprotein B (pCIgB) of pseudorabies virus (PrV) was used to evaluate both the nature of the induced immunity and protection against a virulent virus. With regard to systemic delivery, the intramuscular (i.m.) immunization with pCIgB induced strong PrV-specific IgG responses in serum but was inefficient in generating a mucosal IgA response. Mucosal delivery through intranasal (i.n.) immunization of pCIgB induced both systemic and mucosal immunity at the distal mucosal site. However, the levels of systemic immunity induced by i.n. immunization were less than those induced by i.m. immunization. Moreover, i.n. genetic transfer of pCIgB appeared to induce Th2-biased immunity compared with systemic delivery, as judged by the ratio of PrV-specific IgG isotypes and Th1- and Th2-type cytokines produced by stimulated T cells. Moreover, the immunity induced by i.n. immunization did not provide effective protection against i.n. challenge of a virulent PrV strain, whereas i.m. immunization produced resistance to viral infection. Therefore, although i.n. immunization was a useful route for inducing mucosal immunity at the virus entry site, i.n. immunization did not provide effective protection against the lethal infection of PrV.

Characterization of recombinant foot-and-mouth disease virus pentamer-like structures expressed by baculovirus and their use as diagnostic antigens in a blocking ELISA.

Non-infectious recombinant pentamer-like structures of the foot-and-mouth disease virus (FMDV) were expressed by baculovirus, and the antigenicity and immunogenicity of the proteins were analyzed in a blocking ELISA for the detection of FMDV antibodies. The recombinant pentamer-like structures were produced in insect (Sf9) cells that were inoculated with recombinant baculoviruses that expressed, simultaneously, the genes for the P1 and 3C proteins of FMDV from individual promoters. The FMDV pentamer-like structures were processed by viral 3C protease, as shown in Western blots, and were antigenic, as revealed by their reactivities in an indirect ELISA. Analysis by CsCl gradient centrifugation showed that the pentamer-like structures were similar to authentic pentameric subunits from FMDV in terms of sedimentation velocity. Furthermore, the pentamer-like structures induced high levels of FMDV-specific antibodies in mice following immunization. Observations made under the electron microscope revealed that the pentamer-like structures expressed by insect cells self-assembled to form pentameric subunits of 7-8 nm in diameter, which resemble the authentic FMDV (23+/-2 nm in diameter). The results indicate that these pentamer-like structures are as antigenic and immunogenic as authentic FMDV, although the former are smaller in size. Based on these results, a blocking ELISA was developed using the recombinant pentamer-like structure. The ELISA showed specificity of 99.5% and sensitivity of 98.5% when tested with FMDV antibody-negative and -positive sera, respectively. This blocking ELISA is highly specific and offers many advantages over the current ELISAs that use inactivated FMDV antigen. This is the first report of the production and diagnostic application of recombinant pentameric subunits of FMDV.

Correlation between the nature of immunity induced by different immunogens and the establishment of latent infection by wild-type pseudorabies virus.

To assess the correlation between the nature of immunity induced by different types of immunogens and the establishment of latent infection by wild-type pseudorabies virus (PrV), we used a murine model immunized with different immunogens, the PrV modified live vaccine (MLV), inactivated vaccine (IAV), and commercial oil-adjuvant subunit vaccine (OSV), via either intranasal (i.n.) or intramuscular (i.m.) route. Both MLV and IAV induced a different nature of immunity biased to Th1- and Th2-type, respectively, as judged by the ratio of PrV-specific IgG isotypes (IgG2a/IgG1) and the profile of cytokine IL-2, IL-4, and IFN-gamma production. In contrast, the OSV induced a lower isotype IgG2a to IgG1 ratio and higher level of IL-2 production. The MLV (inducing Th1-type) provided more effective protection against a virulent wild-type PrV challenge than IAV and OSV (inducing Th2- and mixed type, respectively). In addition, the MLV impeded the establishment of a latent infection with wild-type PrV, and the decrease in the PrV latency load by immunization with the MLV appeared to be mediated by the immune T-cells. These results demonstrate the substantial role of the immune responses driven by preceding vaccination in modulating the establishment of PrV latency caused by the post-infection of a field virus.

Systemic and mucosal immunity induced by oral somatic transgene vaccination against glycoprotein B of pseudorabies virus using live attenuated Salmonella typhimurium.

Glycoprotein B mediates the absorption and penetration of the pseudorabies virus in the form of an immunodominant Ag, and represents a major target for the development of new vaccines. This study evaluated the efficiency of live attenuated Salmonella typhimurium SL7207 for the oral delivery of DNA vaccine encoding the pseudorabies virus glycoprotein B (pCI-PrVgB) in vivo, leading to the generation of both systemic and mucosal immunity against the pseudorabies virus Ag. An oral transgene vaccination of pCI-PrVgB using a Salmonella carrier produced a broad spectrum of immunity at both the systemic and mucosal sites, whereas the intramuscular administration of a naked DNA vaccine elicited no mucosal immunoglobulin (Ig)A response. Interestingly, the Salmonella-mediated oral transgene vaccination of the pseudorabies virus glycoprotein B biased the immune responses to the Th2-type, as determined by the IgG2a/IgG1 ratio and the cytokine production profile. However, oral vaccination mediated by Salmonella harbouring pCI-PrVgB showed inferior protection to systemic immunization against virulent pseudorabies virus infection. The expression of transgene delivered by Salmonella bacteria in antigen-presenting cells of both the systemic and mucosal-associated lymphoid tissues was further demonstrated. These results highlight the potential use of live attenuated S. typhimurium for an oral transgene pseudorabies virus glycoprotein B vaccination to induce broad immune responses.

Molecular characterization of recent Korean porcine reproductive and respiratory syndrome (PRRS) viruses and comparison to other Asian PRRS viruses.

Twenty-eight PRRS viruses (PRRSVs) isolated from various pig farms in Korea between 2002 and 2003 were sequenced for open-reading frame (ORF) 5 and/or full-length genome and compared with numerous PRRSVs reported from North America, Europe and Asia. All Korean isolates examined were genetically of the North American genotype. The ORF5 sequence of one isolate was identical to Ingelvac PRRS MLV vaccine virus. ORF5 nucleotide sequence divergence of the remaining 27 Korean PRRSVs from VR-2332, the prototype of the North American PRRSV and parental strain of the MLV vaccine virus, ranged from 1.3% to 12.9%, which corresponded to 2.0% to 14.9% divergence at the amino acid level, raising a concern on the efficacy of the MLV vaccine. Phylogenetic analyses of ORF5 and/or full-length sequences revealed that the Korean PRRSVs formed a clade distinct from PRRSVs reported from other Asian countries (China, Taiwan, Japan, and Thailand). Our study demonstrated that PRRSVs of the North American genotype were introduced to the Korean swine population some time ago and have evolved independently from PRRSV in other Asian countries, suggesting that geographic separation might influence the molecular evolution of PRRSV. This should be taken into consideration when a national PRRS prevention and control policy for international trade is established.

Investigation of pseudorabies virus latency in nervous tissues of seropositive pigs exposed to field strain.

The prevalence and quantity of latent pseudorabies virus (PrV) in nervous tissues of pigs exposed to field strain in Korea was investigated by nested and real-time PCR. Nervous tissues including trigeminal ganglion (TG), olfactory bulb (OB), and brain stem (BS) were collected from 94 seropositive pigs. PrV latent infection in nervous tissues was initially investigated by nested PCR targeting three glycoprotein genes (gB, gE, and gG). Based on the obtained result, latent infection was detected in 95.7% of screened animals. Furthermore, it was revealed that the examined tissues harbored different copy numbers of latent PrV genome ranging from <10(2.0) to 10(7.1) copies per microgram of genomic DNA in real-time PCR analysis. These results show that under normal conditions, levels of latent PrV in the nervous tissues of pigs can vary across a wide range. Therefore, the data presented here provides information regarding control of the endemic state of PrV in Korea.

Development of synthetic peptide ELISA based on nonstructural protein 2C of foot and mouth disease virus.

It was reported that the sera of convalescent animals contain antibodies to foot and mouth disease (FMD) virus (FMDV) 2C, highly conserved nonstructural protein (NSP), whereas the sera of vaccinated animals do not. But ELISA methods using this protein were not reported and developed until recently. In this study, NSP 2C peptides were synthesized within the amino acid sequence of the conserved 2C nonstructural region of FMDV according to the sequences from Genbank database and used for identifying antigenic determinants. One of the synthesized thirteen peptides gave strong positive reactivity with most of the sera from 13 FMD infected farms, but not with sera from vaccinated and non-infected animals. Moreover, with the sera collected through serial bleedings from four cattle and five goats infected with FMDV O/SKR/2000 experimentally, positive results were obtained in two species after 10 days post infection (DPI). Therefore, we tried to develop and evaluate this ELISA based on 2C peptides. In comparison with the commercial NSP ELISA, the 2C peptide based ELISA method showed good specificity and sensitivity. These results demonstrate that the synthetic 2C peptide ELISA can be a complementary marker to differentiate FMDV-infected from vaccinated on a herd basis.

Molecular survey of latent pseudorabies virus infection in nervous tissues of slaughtered pigs by nested and real-time PCR.

In this study, the prevalence and quantity of a latent pseudorabies virus (PrV) infection in the nervous tissues of randomly selected pigs was determined via nested and real-time PCR. The nervous tissues, including the trigeminal ganglion (TG), olfactory bulb (OB), and brain stem (BS), were collected from the heads of 40 randomly selected pigs. The majority of the nervous tissues from the selected pigs evidenced a positively amplified band on nested PCR. In particular, nested PCR targeted to the PrV glycoprotein B (gB) gene yielded positive results in all of the BS samples. Nested PCR for either the gE or gG gene produced positive bands in a less number of nervous tissues (57.5% and 42.5%, respectively). Real-time PCR revealed that the examined tissues harbored large copy numbers of latent PrV DNA, ranging between 10(0.1) and 10(7.2) (1-1.58 x 10(7)) copies per 1 microg of genomic DNA. Real-time PCR targeted to the PrV gE gene exhibited an accumulated fluorescence of reporter dye at levels above threshold, thereby indicating a higher prevalence than was observed on the nested PCR (100% for BS, 92% for OB, and 85% for TG). These results indicate that a large number of farm-grown pigs are latently infected with a field PrV strain with a variety of copy numbers. This result is similar to what was found in association with the human herpes virus.

Development of a Lightcycler-based reverse transcription polymerase chain reaction for the detection of foot-and-mouth disease virus.

One step TaqMan real-time reverse transcription polymerase chain reaction (R/T RT-PCR) using a set of primers/probes was developed for the detection of foot-and-mouth disease (FMD) virus. The gene-specific probes labeled fluorogen for the internal ribosomal entry site, Leader sequence and 2B regions were used to detect FMD virus (FMDV). This assay specifically detected FMDV both in cell culture preparations and clinical samples, and was capable of distinguishing FMD from other viral diseases similar to clinical signs (swine vesicular disease, vesicular stomatitis and bovine viral diarrhea). This assay was shown to be 1000-fold more sensitive than the conventional RT-PCR method. The detection limits of this assay was 1 TCID(50)/ml of the FMDV RNA concentration. Quantification was obtained by a standard curves plotting threshold cycle values versus known infectivity titer. The assay was sensitive, specific and rapid enough to detect FMDV RNA genome in probang samples. As such, the described method is reliable and provides faster disease diagnostics than the conventional RT-PCR procedure to detect FMDV.

Identification and antigenic site analysis of foot-and-mouth disease virus from pigs and cattle in Korea.

From May to June 2002, a total of 16 foot-and mouth disease (FMD) outbreaks due to the serotype O virus, Pan Asia strain, were recorded in Korea. The viruses were identified by antigen ELISA, RT-PCR and sequence analysis. The overall nucleotide sequence divergence of the VP1 region among the 4 isolates in 2002 was 0 to 1.4%, but between O/SKR/2002 and O/SKR/2000 isolates was 1.9-4.9%. Phylogenetic analysis with the some known strains from East Asian countries showed that the 4 Korean isolates in 2002 formed one distinct cluster, which different from clusters of Korean isolates in 2000, with in the same lineage of the ME-SA topotype strains. Deduced amino acid sequences around neutralizable antigenic site on VP1 site of O/SKR/2002 isolates were aligned and compared with other strains. At the antigenic site 1, the replacements of the critical amino acid residues at position 144 from V to L and at position 152 from A to T were observed in O/SKR/2002 viruses. For antigenic site 2 and 4, there were not significant variations in general. At the antigenic site 3, the substitutions of amino acid residues were present at positions 54 and 56 in O/SKR/2002 isolates and an alternative residue I at position 54 are observed only at the sequence of O/SKR/AS/2002 (cow) virus. And the substitution (L-->P) of significant residue at position 144 was detected at the amino acid sequence of the O/SKR/2002 (cow) virus.