Detection and disease diagnosis trends (2017-2022) for Streptococcus suis, Glaesserella parasuis, Mycoplasma hyorhinis, Actinobacillus suis and Mycoplasma hyosynoviae at Iowa State University Veterinary Diagnostic Laboratory.

BACKGROUND: Accurate measurement of disease associated with endemic bacterial agents in pig populations is challenging due to their commensal ecology, the lack of disease-specific antemortem diagnostic tests, and the polymicrobial nature of swine diagnostic cases. The main objective of this retrospective study was to estimate temporal patterns of agent detection and disease diagnosis for five endemic bacteria that can cause systemic disease in porcine tissue specimens submitted to the Iowa State University Veterinary Diagnostic Laboratory (ISU VDL) from 2017 to 2022. The study also explored the diagnostic value of specific tissue specimens for disease diagnosis, estimated the frequency of polymicrobial diagnosis, and evaluated the association between phase of pig production and disease diagnosis. RESULTS: S. suis and G. parasuis bronchopneumonia increased on average 6 and 4.3%, while S. suis endocarditis increased by 23% per year, respectively. M. hyorhinis and A. suis associated serositis increased yearly by 4.2 and 12.8%, respectively. A significant upward trend in M. hyorhinis arthritis cases was also observed. In contrast, M. hyosynoviae arthritis cases decreased by 33% average/year. Investigation into the diagnostic value of tissues showed that lungs were the most frequently submitted sample, However, the use of lung for systemic disease diagnosis requires caution due to the commensal nature of these agents in the respiratory system, compared to systemic sites that diagnosticians typically target. This study also explored associations between phase of production and specific diseases caused by each agent, showcasing the role of S. suis arthritis in suckling pigs, meningitis in early nursery and endocarditis in growing pigs, and the role of G. parasuis, A. suis, M. hyorhinis and M. hyosynoviae disease mainly in post-weaning phases. Finally, this study highlighted the high frequency of co-detection and -disease diagnosis with other infectious etiologies, such as PRRSV and IAV, demonstrating that to minimize the health impact of these endemic bacterial agents it is imperative to establish effective viral control programs. CONCLUSIONS: Results from this retrospective study demonstrated significant increases in disease diagnosis for S. suis, G. parasuis, M. hyorhinis, and A. suis, and a significant decrease in detection and disease diagnosis of M. hyosynoviae. High frequencies of interactions between these endemic agents and with viral pathogens was also demonstrated. Consequently, improved control programs are needed to mitigate the adverse effect of these endemic bacterial agents on swine health and wellbeing. This includes improving diagnostic procedures, developing more effective vaccine products, fine-tuning antimicrobial approaches, and managing viral co-infections.

Diagnosis of Anaplasma marginale in cattle at the Iowa State University veterinary diagnostic laboratory 2003-2021.

Anaplasma marginale is a blood-borne rickettsia-like organism that infects cattle erythrocytes and causes anaplasmosis. This study reviews diagnostic data of all A. marginale diagnostics performed from 2003 to August 2021 in the Iowa State Veterinary Diagnostic Laboratory. Typically, the referring veterinarian's initial tentative diagnosis was based on presenting clinical signs or necropsy findings. Confirmatory testing at the ISU-VDL consisted of light microscopy evaluation of stained blood smears or molecular diagnostic procedures. A total of 94 cases were submitted with tissue samples from deceased animals, of which 79 were from Iowa and 15 were from other states. The most typical gross lesions were widespread yellow adipose tissue and splenomegaly. Typical histopathological lesions included marked bile stasis and hemosiderin-laden macrophages in the liver and spleen, respectively. Starting in 2013, when PCR was implemented to confirm cases of anaplasmosis, 315/1125 (28%) were positive to A. marginale, and 810 were negative, using a cut-off of 35.0 Ct. The average (±SD) of the positive PCR Ct was 19.5 (±6.0), and the first and third quartiles were 14.9 and 23.4. Most cases occurred between August and November, peaking in September, whether from necropsies or positive blood samples by PCR. The most common tick observed in Iowa, Dermacentor variabilis, is likely the main vector for transmission. Further surveys should be conducted to estimate seroprevalence by geographical location, the density of cattle populations, distribution of known vectors according to season, and strains of A. marginale.

Utilizing productivity and health breeding-to-market information along with disease diagnostic data to identify pig mortality risk factors in a U.S. swine production system.

Aggregated diagnostic data collected over time from swine production systems is an important data source to investigate swine productivity and health, especially when combined with records concerning the pre-weaning and post-weaning phases of production. The combination of multiple data streams collected over the lifetime of the pigs is the essence of the whole-herd epidemiological investigation. This approach is particularly valuable for investigating the multifaceted and ever-changing factors contributing to wean-to-finish (W2F) swine mortality. The objective of this study was to use a retrospective dataset ("master table") containing information on 1,742 groups of pigs marketed over time to identify the major risk factors associated with W2F mortality. The master table was built by combining historical breed-to-market performance and health data with disease diagnostic records (Dx Codes) from marketed groups of growing pigs. After building the master table, univariate analyses were conducted to screen for risk factors to be included in the initial multivariable model. After a stepwise backward model selection approach, 5 variables and 2 interactions remained in the final model. Notably, the diagnosis variable significantly associated with W2F mortality was porcine reproductive and respiratory syndrome virus (PRRSV). Closeouts with clinical signs suggestive of Salmonella spp. or Escherichia coli infection were also associated with higher W2F mortality. Source sow farm factors that remained significantly associated with W2F mortality were the sow farm PRRS status, average weaning age, and the average pre-weaning mortality. After testing for the possible interactions in the final model, two interactions were significantly associated with wean-to-finish pig mortality: (1) sow farm PRRS status and a laboratory diagnosis of PRRSV and (2) average weaning age and a laboratory diagnosis of PRRS. Closeouts originating from PRRS epidemic or PRRS negative sow farms, when diagnosed with PRRS in the growing phase, had the highest W2F mortality rates. Likewise, PRRS diagnosis in the growing phase was an important factor in mortality, regardless of the average weaning age of the closeouts. Overall, this study demonstrated the utility of a whole-herd approach when analyzing diagnostic information along with breeding-to-market productivity and health information, to measure the major risk factors associated with W2F mortality in specified time frames and pig populations.

Salmonella enterica serovar Brandenburg abortions in dairy cattle.

Two separate late-term abortion outbreaks in Jersey heifers in July 2020 and December 2020 were investigated by the Iowa State University Veterinary Diagnostic Laboratory. We evaluated 3 whole fetuses and 11 sets of fresh and formalin-fixed fetal tissues during the course of the outbreaks. The late-term abortions were first identified at a heifer development site and subsequently observed at the dairy farm. Aborted fetuses had moderate-to-marked postmortem autolysis with no gross lesions identified. Observed clinical signs in cows at the dairy farm ranged from intermittent loose stools to acute post-abortion pyrexia and reduced feed intake. Routine histopathology and reproductive bacterial culture revealed acute, suppurative placentitis with moderate-to-heavy growth of Salmonella spp. group B from stomach contents, liver, placenta, and heifer fecal contents. Serotyping identified Salmonella enterica subsp. enterica serovar Brandenburg in all 14 fresh tissue cases, as well as individual and pooled heifer feces. Whole-genome sequencing analysis revealed that all isolates belonged to ST type 873 and possessed typhoid toxin genes, several fimbrial gene clusters, type III secretion system genes, and several pathogenicity islands. Abortions caused by Salmonella Brandenburg have not been reported previously in dairy cattle in the United States, to our knowledge.

Bovine coronavirus in the lower respiratory tract of cattle with respiratory disease.

Bovine coronavirus (BCoV) is a known cause of enteric disease in cattle; however, its role in bovine respiratory disease (BRD) is poorly understood, with a dearth of evidence of the detection of the virus in respiratory tract lesions. We coupled histologic evaluation of tracheal and lower airway tissues from 104 calves with BRD in which BCoV was detected in the lungs via PCR followed by direct detection of BCoV by immunohistochemistry and an RNA in situ hybridization assay (ISH; RNAscope technology). RNAscope ISH detected BCoV in respiratory epithelium in more cases than did IHC. Using both methods of direct detection, tracheal epithelial attenuation and identification of the virus within lesions were observed commonly. Our results confirm a role of BCoV in respiratory tract infection and pathology, and show that the virus likely plays a role in the development of BRD.

Streptococcus gallolyticus and Bacterial Endocarditis in Swine, United States, 2015-2020.

To evaluate trends in bacterial causes of valvular endocarditis in swine, we retrospectively analyzed 321 cases diagnosed at Iowa State University Veterinary Diagnostic Laboratory (Ames, IA, USA) during May 2015--April 2020. Streptococcus gallolyticus was the causative agent for 7.59% of cases. This emerging infection in swine could aid study of endocarditis in humans.

Distribution and persistence of atypical porcine pestivirus in experimentally inoculated pigs.

Atypical porcine pestivirus (APPV) is a cause of congenital tremors (CTs) in piglets and has been found in swine populations around the globe. Although systemic distribution of the virus has been reported, there is limited information regarding viral localization at the cellular level and distribution at the tissue level. We collected multiple tissues from 2-d-old piglets (n = 36) born to sows inoculated at 45 or 62 d of gestation with APPV via 3 simultaneous routes: intravenous, intranasal, and directly in amniotic vesicles. In addition, 2 boars from APPV-inoculated sows with CT were raised and euthanized when 11 mo old. In situ hybridization performed on tissue samples from piglets demonstrated a broad and systemic distribution of viral RNA including endothelial cells, fibroblasts, and smooth muscle. Labeling in tissues was more pronounced in piglet tissues compared to boars, with the notable exception of diffuse labeling of the cerebellum in boars. Presence of APPV in boar tissues well after resolution of clinical signs suggests persistence of APPV similar to other pestiviruses.

Visualization and application of disease diagnosis codes for population health management using porcine diseases as a model.

Accurate and timely results of diagnostic investigations and laboratory testing guide clinical interventions for the continuous improvement of animal health and welfare. Infectious diseases can severely limit the health, welfare, and productivity of populations of animals. Livestock veterinarians submit thousands of samples daily to veterinary diagnostic laboratories (VDLs) for disease diagnosis, pathogen monitoring, and surveillance. Individual diagnostic laboratory reports are immediately useful; however, aggregated historical laboratory data are increasingly valued by clinicians and decision-makers to identify changes in the health status of various animal populations over time and geographical space. The value of this historical information is enhanced by visualization of trends of agent detection, disease diagnosis, or both, which helps focus time and resources on the most significant pathogens and fosters more effective communication between livestock producers, veterinarians, and VDL professionals. Advances in data visualization tools allow quick, efficient, and often real-time scanning and analysis of databases to inform, guide, and modify animal health intervention algorithms. Value is derived at the farm, production system, or regional level. Visualization tools allow client-specific analyses, benchmarking, formulation of research questions, and monitoring the effects of disease management and precision farming practices. We present here the approach taken to visualize trends of disease occurrence using porcine disease diagnostic code data for the period 2010 to 2019. Our semi-automatic standardized creation of a visualization platform allowed the transformation of diagnostic report data into aggregated information to visualize and monitor disease diagnosis.

Disease diagnostic coding to facilitate evidence-based medicine: current and future perspectives.

Technologic advances in information management have rapidly changed laboratory testing and the practice of veterinary medicine. Timely and strategic sampling, same-day assays, and 24-h access to laboratory results allow for rapid implementation of intervention and treatment protocols. Although agent detection and monitoring systems have progressed, and wider tracking of diseases across veterinary diagnostic laboratories exists, such as by the National Animal Health Laboratory Network (NAHLN), the distinction between detection of agent and manifestation of disease is critical to improved disease management. The implementation of a consistent, intuitive, and useful disease diagnosis coding system, specific for veterinary medicine and applicable to multiple animal species within and between veterinary diagnostic laboratories, is the first phase of disease data aggregation. Feedback loops for continuous improvement that could aggregate existing clinical and laboratory databases to improve the value and applications of diagnostic processes and clinical interventions, with interactive capabilities between clinicians and diagnosticians, and that differentiate disease causation from mere agent detection, remain incomplete. Creating an interface that allows aggregation of existing data from clinicians, including final diagnosis, interventions, or treatments applied, and measures of outcomes, is the second phase. Prototypes for stakeholder cooperation, collaboration, and beta testing of this vision are in development and becoming a reality. We focus here on how such a system is being developed and utilized at the Iowa State University Veterinary Diagnostic Laboratory to facilitate evidence-based medicine and utilize diagnostic coding for continuous improvement of animal health and welfare.

Data standardization implementation and applications within and among diagnostic laboratories: integrating and monitoring enteric coronaviruses.

Every day, thousands of samples from diverse populations of animals are submitted to veterinary diagnostic laboratories (VDLs) for testing. Each VDL has its own laboratory information management system (LIMS), with processes and procedures to capture submission information, perform laboratory tests, define the boundaries of test results (i.e., positive or negative), and report results, in addition to internal business and accounting applications. Enormous quantities of data are accumulated and stored within VDL LIMSs. There is a need for platforms that allow VDLs to exchange and share portions of laboratory data using standardized, reliable, and sustainable information technology processes. Here we report concepts and applications for standardization and aggregation of data from swine submissions to multiple VDLs to detect and monitor porcine enteric coronaviruses by RT-PCR. Oral fluids, feces, and fecal swabs were the specimens submitted most frequently for enteric coronavirus testing. Statistical algorithms were used successfully to scan and monitor the overall and state-specific percentage of positive submissions. Major findings revealed a consistently recurrent seasonal pattern, with the highest percentage of positive submissions detected during December-February for porcine epidemic diarrhea virus, porcine deltacoronavirus, and transmissible gastroenteritis virus (TGEV). After 2014, very few submissions tested positive for TGEV. Monitoring VDL data proactively has the potential to signal and alert stakeholders early of significant changes from expected detection. We demonstrate the importance of, and applications for, data organized and aggregated by using LOINC and SNOMED CTs, as well as the use of customized messaging to allow inter-VDL exchange of information.

Case Report and Genomic Characterization of a Novel Porcine Nodavirus in the United States.

Nodaviruses are small bisegmented RNA viruses belonging to the family Nodaviridae. Nodaviruses have been identified in different hosts, including insects, fishes, shrimps, prawns, dogs, and bats. A novel porcine nodavirus was first identified in the United States by applying next-generation sequencing on brain tissues of pigs with neurological signs, including uncontrollable shaking. RNA1 of the porcine nodavirus had the highest nucleotide identity (51.1%) to the Flock House virus, whereas its RNA2 shared the highest nucleotide identity (48%) with the RNA2 segment of caninovirus (Canine nodavirus). Genetic characterization classified porcine nodavirus as a new species under the genus Alphanodavirus. Further studies are needed to understand the pathogenicity and clinical impacts of this virus.

Cases of high mortality in cull sows and feeder pigs associated with Streptococcus equi subsp. zooepidemicus septicemia.

Investigations of 2 cases of high mortality in cull sows and feeder pigs from a buying station in Ohio and cull sows at an abattoir in Tennessee were conducted at the Iowa State University Veterinary Diagnostic Laboratory. The animals were presented as weak, lethargic, and some with high fever. Rapidly escalating mortality was reported to be as high as 30-50% within groups at the buying station over 8-10 d, and 30-40% over 5-7 d at the abattoir. Splenomegaly and red lymph nodes were the most consistent macroscopic findings, with scant fibrinous polyserositis observed in one sow. The microscopic lesions of vasculitis, fibrin thrombi, fibrinosuppurative polyserositis, and intralesional bacteria were consistent with acute bacterial septicemia. Bacterial culture isolated Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) from multiple organs, including spleen, lung, and kidney. PCR tests were negative for African swine fever virus, classical swine fever virus, Erysipelothrix rhusiopathiae, porcine reproductive and respiratory syndrome virus, porcine circovirus 2, and Salmonella spp. Porcine circovirus 3 was inconsistently detected at low levels by PCR, with a lack of associated lesions. Next-generation sequencing identified S. zooepidemicus and porcine partetravirus in the serum sample of the feeder pig from the buying station. Phylogenetic analysis of the szP gene indicated that the S. zooepidemicus isolates from Ohio and Tennessee are in genotype VI. We conclude that the cause of these high mortality events in swine was S. zooepidemicus septicemia.

Genetic characterization of Streptococcus equi subspecies zooepidemicus associated with high swine mortality in the United States.

High mortality events due to Streptococcus equi subspecies zooepidemicus (Streptococcus zooepidemicus) in swine have not previously been reported in the United States. In September and October 2019, outbreaks with swine mortality up to 50% due to S. zooepidemicus septicaemia were reported in Ohio and Tennessee. Genomic epidemiological analysis revealed that the eight outbreak isolates were clustered together with ATCC 35246, a Chinese strain caused outbreaks with high mortality, also closely related to three isolates from human cases from Virginia, but significantly different from an outbreak-unrelated swine isolate from Arizona and most isolates from other animal species. Comparative genomic analysis on two outbreak isolates and another outbreak-unrelated isolate identified several genomic islands and virulence genes specifically in the outbreak isolates only, which are likely associated with the high mortality observed in the swine population. These findings have implications for understanding, tracking and possibly preventing diseases caused by S. zooepidemicus in swine.

Prediction of seasonal patterns of porcine reproductive and respiratory syndrome virus RNA detection in the U.S. swine industry.

We developed a model to predict the cyclic pattern of porcine reproductive and respiratory syndrome virus (PRRSV) RNA detection by reverse-transcription real-time PCR (RT-rtPCR) from 4 major swine-centric veterinary diagnostic laboratories (VDLs) in the United States and to use historical data to forecast the upcoming year's weekly percentage of positive submissions and issue outbreak signals when the pattern of detection was not as expected. Standardized submission data and test results were used. Historical data (2015-2017) composed of the weekly percentage of PCR-positive submissions were used to fit a cyclic robust regression model. The findings were used to forecast the expected weekly percentage of PCR-positive submissions, with a 95% confidence interval (CI), for 2018. During 2018, the proportion of PRRSV-positive submissions crossed 95% CI boundaries at week 2, 14-25, and 48. The relatively higher detection on week 2 and 48 were mostly from submissions containing samples from wean-to-market pigs, and for week 14-25 originated mostly from samples from adult/sow farms. There was a recurring yearly pattern of detection, wherein an increased proportion of PRRSV RNA detection in submissions originating from wean-to-finish farms was followed by increased detection in samples from adult/sow farms. Results from the model described herein confirm the seasonal cyclic pattern of PRRSV detection using test results consolidated from 4 VDLs. Wave crests occurred consistently during winter, and wave troughs occurred consistently during the summer months. Our model was able to correctly identify statistically significant outbreak signals in PRRSV RNA detection at 3 instances during 2018.

Increasing the ratio of SID lysine to metabolizable energy improves pig performance during a viral challenge.

Porcine reproductive and respiratory syndrome virus (PRRSV) compromises pig performance. However, increasing standardized ileal digestible Lys per Mcal metabolizable energy (SID Lys:ME) above requirement has been shown to mitigate reduced performance seen during a porcine reproductive and respiratory syndrome (PRRS) virus challenge. The objective of this study was to evaluate the effects of increasing the dietary SID Lys:ME from 100% National Research Council (NRC) requirement to 120% of the requirement in vaccinated (vac+; modified live vaccine Ingelvac PRRS) and non-vaccinated (vac-; no PRRS vaccine) grower pigs subjected to a PRRSV challenge. In addition, the dietary formulation approach to achieve the 120% ratio by increasing Lys relative to energy (HL) or diluting energy in relation to Lys (LE) was evaluated. This allowed us to test the hypothesis that pigs undergoing a health challenge would have the ability to eat to their energy needs. Within vaccine status, 195 mixed-sex pigs, vac+ (35.2 ± 0.60 kg body weight [BW]) and vac- (35.2 ± 0.65 kg BW) were randomly allotted to one of three dietary treatments (2.67, 3.23, or 3.22 g SID Lys:ME) for a 42-d PRRS virus challenge study representing 100%, 120%, and 120% of NRC requirement, respectively. Pigs were randomly allotted across two barns, each containing 24 pens with 7 to 10 pigs per pen (8 pens per diet per vaccine status). On day post-inoculation 0, both barns were inoculated with PRRSV and started on experimental diets. Within vaccine status, weekly and overall challenge period pig performance were assessed. In both vac+ (P < 0.05) and vac- (P < 0.05) pigs, the HL and LE diets increased end BW and overall average daily gain (ADG) ADG compared with pigs fed the control diet (P < 0.05). Overall, average daily feed intake (ADFI) during the challenge period was greater (P < 0.05) for pigs fed the LE diet compared with pigs fed control and HL treatments, regardless of vaccine status (20% and 17% higher ADFI than the control in vac+ and vac- pigs, respectively). The HL vac+ pigs had the greatest gain to feed (G:F) compared with the control and LE pigs (0.438 vs. 0.394 and 0.391 kg/kg, respectively; P < 0.01). Feed efficiency was not impacted (P > 0.10) by treatment in the vac- pigs. In summary, PRRSV-challenged grower pigs consumed feed to meet their energy needs as indicated by the increase in ADFI when energy was diluted in the (LE) diet, compared with control pigs. In both PRRS vac+ and vac- pigs subsequently challenged with PRRSV, regardless of formulation approach, fed 120% SID Lys:ME diets resulted in enhanced overall growth performance.

Isolation of PCV3 from Perinatal and Reproductive Cases of PCV3-Associated Disease and In Vivo Characterization of PCV3 Replication in CD/CD Growing Pigs.

Porcine circovirus 3 (PCV3) has been identified as a putative swine pathogen with a subset of infections resulting in stillborn and mummified fetuses, encephalitis and myocarditis in perinatal, and periarteritis in growing pigs. Three PCV3 isolates were isolated from weak-born piglets or elevated stillborn and mummified fetuses. Full-length genome sequences from different passages and isolates (PCV3a1 ISU27734, PCV3a2 ISU58312, PCV3c ISU44806) were determined using metagenomics sequencing. Virus production in cell culture was confirmed by qPCR, IFA, and in situ hybridization. In vivo replication of PCV3 was also demonstrated in CD/CD pigs (n = 8) under experimental conditions. Viremia, first detected at 7 dpi, was detected in all pigs by 28 dpi. IgM antibody response was detected between 7-14 dpi in 5/8 PCV3-inoculated pigs but no IgG seroconversion was detected throughout the study. Pigs presented histological lesion consistent with multi systemic inflammation characterized by myocarditis and systemic perivasculitis. Viral replication was confirmed in all tissues by in situ hybridization. Clinically, all animals were unremarkable throughout the study. Although the clinical relevance of PCV3 remains under debate, this is the first isolation of PCV3 from perinatal and reproductive cases of PCV3-associated disease and in vivo characterization of PCV3 infection in a CD/CD pig model.

Polioencephalomyelitis in Domestic Swine Associated With Porcine Astrovirus Type 3.

In the past decade, different members of the genus Mamastrovirus have been associated with outbreaks of neurologic disease in humans, cattle, sheep, mink, and, most recently, porcine astrovirus 3 (PoAstV3) in swine. We performed a retrospective analysis of 50 cases of porcine neurologic disease of undetermined cause but with microscopic lesions compatible with a viral encephalomyelitis to better understand the role and pathogenesis of PoAstV3 infection. Nucleic acid was extracted from formalin-fixed paraffin-embedded (FFPE) tissue for reverse transcription quantitative polymerase chain reaction (RT-qPCR) testing for PoAstV3. In addition, 3 cases with confirmed PoAstV3-associated disease were assayed by RT-qPCR to investigate PoAstV3 tissue distribution. PoAstV3 was detected in central nervous system (CNS) tissue via RT-qPCR and in situ hybridization in 13 of 50 (26%) FFPE cases assayed. PoAstV3 was rarely detected in any tissues outside the CNS. Positive cases from the retrospective study included pigs in various production categories beginning in 2010, the earliest year samples were available. Based on these results, PoAstV3 appears to be a recurring putative cause of viral encephalomyelitis in swine that is rarely detected outside of the CNS at the time of clinical neurologic disease, unlike other common viral causes of neurologic disease in swine.

Emergence of Salmonella enterica serovar 4,[5],12:i:- as the primary serovar identified from swine clinical samples and development of a multiplex real-time PCR for improved Salmonella serovar-level identification.

Rapid identification of the infecting Salmonella serovar from porcine diagnostic samples is vital to allow implementation of appropriate on-farm treatment and management decisions. Although identification at the serogroup level can be rapidly achieved at most veterinary diagnostic laboratories, final Salmonella serovar identification often takes several weeks because of the limited number of reference laboratories performing the complex task of serotyping. Salmonella serogroup B, currently the dominant serogroup identified from swine clinical samples in the United States, contains serovars that vary from highly pathogenic to minimally pathogenic in swine. We determined the frequency of detection of individual group B serovars at the Iowa State Veterinary Diagnostic Laboratory from 2008 to 2017, and validated a multiplex real-time PCR (rtPCR) to distinguish pathogenic serogroup B serovars from those of lesser pathogenicity. Our results indicate that, since 2014, Salmonella enterica ssp. enterica serovar 4,[5],12:i:- has been the dominant serovar identified from swine clinical samples at the ISU-VDL, with S. Typhimurium now the second most common serovar identified. We developed a rtPCR to allow rapid differentiation of samples containing S. 4,[5],12:i:- and S. Typhimurium from samples containing serovars believed to be of less pathogenicity, such as S. Agona and S. Derby. When combined with enrichment culture, this rtPCR has the ability to significantly improve the time to final serovar identification of the 2 most commonly identified pathogenic Salmonella serovars in swine, and allows rapid implementation of serovar-specific intervention strategies.

Macroepidemiological aspects of porcine reproductive and respiratory syndrome virus detection by major United States veterinary diagnostic laboratories over time, age group, and specimen.

This project investigates the macroepidemiological aspects of porcine reproductive and respiratory syndrome virus (PRRSV) RNA detection by veterinary diagnostic laboratories (VDLs) for the period 2007 through 2018. Standardized submission data and PRRSV real-time reverse-transcriptase polymerase chain reaction (RT-qPCR) test results from porcine samples were retrieved from four VDLs representing 95% of all swine samples tested in NAHLN laboratories in the US. Anonymized data were retrieved and organized at the case level using SAS (SAS® Version 9.4, SAS® Institute, Inc., Cary, NC) with the use of PROC DATA, PROC MERGE, and PROC SQL scripts. The final aggregated and anonymized dataset comprised of 547,873 unique cases was uploaded to Power Business Intelligence-Power BI® (Microsoft Corporation, Redmond, Washington) to construct dynamic charts. The number of cases tested for PRRSV doubled from 2010 to 2018, with that increase mainly driven by samples typically used for monitoring purposes rather than diagnosis of disease. Apparent seasonal trends for the frequency of PRRSV detection were consistently observed with a higher percentage of positive cases occurring during fall or winter months and lower during summer months, perhaps due to increased testing associated with well-known seasonal occurrence of swine respiratory disease. PRRSV type 2, also known as North American genotype, accounted for 94.76% of all positive cases and was distributed across the US. PRRSV type 1, also known as European genotype, was geographically restricted and accounted for 2.15% of all positive cases. Co-detection of both strains accounted for 3.09% of the positive cases. Both oral fluid and processing fluid samples, had a rapid increase in the number of submissions soon after they were described in 2008 and 2017, respectively, suggesting rapid adoption of these specimens by the US swine industry for PRRSV monitoring in swine populations. As part of this project, a bio-informatics tool defined as Swine Disease Reporting System (SDRS) was developed. This tool has real-time capability to inform the US swine industry on the macroepidemiological aspects of PRRSV detection, and is easily adaptable for other analytes relevant to the swine industry.

Early detection and differential serodiagnosis of Mycoplasma hyorhinis and Mycoplasma hyosynoviae infections under experimental conditions.

Mycoplasma hyorhinis (MHR) and Mycoplasma hyosynoviae (MHS) are common opportunistic pathogens in the upper respiratory tract and tonsils of swine. The identification of the specific species involved in clinical cases using conventional diagnostic methods is challenging. Therefore, a recombinant chimeric polypeptide based on the seven known variable lipoproteins (A-G) specific of MHR and a cocktail of surface proteins detergent-extracted from MHS cultures were generated and their suitability as antemortem biomarkers for serodiagnosis of MHR- and MHS-infection were evaluated by ELISA. M. hyorhinis and MHS ELISA performance, evaluated using serum samples collected over a 56-day observation period from pigs inoculated with MHR, MHS, M. hyopneumoniae, M. flocculare, or Friis medium, varied by assay, targeted antibody isotype, and cutoffs. The progressions of MHR and MHS clinical diseases were evaluated in relation to the kinetics of the isotype-specific antibody response in serum and bacterial shedding in oral fluids during the observation period. In pigs inoculated with MHR, bacterial DNA was detected in one or more of the 5 pens at all sampling points throughout the study, IgA was first detected at DPI 7, one week before the first clinical signs, with both IgA and IgG detected in all samples collected after DPI 14. The peak of MHS shedding (DPI 8) coincided with the onset of the clinical signs, with both IgA and IgG detected in all serum samples collected ≥ DPI 14. This study demonstrated, under experimental conditions, that both ELISAs were suitable for early detection of specific antibodies against MHR or MHS. The diagnostic performance of the MHR and MHS ELISAs varied depending on the selected cutoff and the antibody isotype evaluated. The high diagnostic and analytical specificity of the ELISAs was particularly remarkable. This study also provides insights into the infection dynamics of MHR-associated disease and MHS-associated arthritis not previously described.