Molecular characterization of the integrative and conjugative elements harbouring multidrug resistance genes in Glaesserella parasuis.

It is widely known that integrative and conjugative elements (ICEs) play an important role in the transmission of resistance genes and other exogenous genes. The present study aimed to characterize the three novel ICEs including ICEGpa76, ICEGpa44, and ICEGpa11, from Glaesserella parasuis. The ICEs from G. parasuis strains d76, Z44, and XP11 were predicted and identified by whole-genome sequencing (WGS) analysis, ICEfinder, and PCR. Characterization of G. parasuis strains carrying ICEs were determined by conjugation assay, antimicrobial susceptibility testing, WGS, phylogenetic analysis, and comparative sequence analysis.The WGS results showed that three ICEs from G. parasuis have a common genetic backbone belonging to characteristics ofthe ICEHpa1 family. The sequence comparison showed that the ICEHpa1 family has five hot spots (HSs) determined by IS6, IS110, and IS256. Moreover, two variable regions (VRs), VR1 and VR2 were determined by multidrug resistance genes and the rearrangement hotspot (rhs) family, respectively. VR1 consists of multidrug resistance genes, ISApl1s, and other accessory genes, while VR2 is composed of IS4, rhs family, transposase, and hypothetical protein genes. Conjugation experiments and MICs revealed that three ICEs could be transferred to G. parasuis strain IV52, indicating these three ICEs could be transmitted horizontally among G. parasuis strains. Additionally, the difference in resistance genes from ICEs might be due to the insertion function of the ISApl1s in VR1, and the rhs family in VR2 might evolve andthen be stably inherited in G. parasuis. These results further elucidated the transmission mechanism of exogenous genes in G. parasuis.

Molecular Characterization and Phylogenetic Analysis of Outer membrane protein P2 (OmpP2) of Glaesserella (Haemophilus) parasuis Isolates in Central State of Peninsular Malaysia.

Glaesserella (Haemophilus) parasuis, the etiological agent of Glässer's disease, is an economically significant pathogen commonly associated with serofibrinous polyserositis, arthritis, fibrinous bronchopneumonia and/or meningitis. This study is the first attempt to molecularly characterize and provide a detailed overview of the genetic variants of G. parasuis present in Malaysia, in reference to its serotype, virulence-associated trimeric autotransporters (vtaA) gene and outer membrane protein P2 (OmpP2) gene. The G. parasuis isolates (n = 11) from clinically sick field samples collected from two major pig producing states (Selangor and Perak) were selected for analysis. Upon multiplex PCR, the majority of the isolates (eight out of 11) were identified to be serotype 5 or 12, and interestingly, serotypes 3, 8 and 15 were also detected, which had never been reported in Malaysia prior to this. Generally, virulent vtaA was detected for all isolates, except for one, which displayed a nonvirulent vtaA. A phylogenetic analysis of the OmpP2 gene revealed that the majority of Malaysian isolates were clustered into genotype 1, which could be further divided into Ia and Ib, while only one isolate was clustered into genotype 2.

Investigation and analysis of etiology associated with porcine respiratory disease complex in China from 2017 to 2021.

Porcine respiratory diseases complex (PRDC) is a highly serious threat to the pig industry. In the present study, we investigated and analyzed the etiology associated with PRDC and explored the role of viruses in respiratory bacterial infections. From 2017 to 2021, clinical samples were collected from 1,307 pigs with typical respiratory symptoms in 269 farms in China and screened for pathogens related to PRDC by PCR and bacterial isolation. The results indicated that PRRSV (41.16%, 95%CI: 38.49~43.83%), PCV2 (21.58%,95%CI: 19.35~23.81%), S. suis (63.50%, 95%CI: 60.89~66.11%), and G. parasuis (28.54%, 95%CI: 26.09~30.99%) were the most commonly detected pathogens in pigs with PRDC in China. The dominant epidemic serotypes (serogroups) of S. suis, G. parasuis, and P. multocida were serotype 2, serotype 1, and capsular serogroups D, respectively. Pigs of different ages exhibited different susceptibilities to these pathogens, e.g., PRRSV, PCV2, and G. parasuis had the highest detection rates in nursery pigs, whereas fattening pigs had the highest detection rates of P. multocida and A. pleuropneumoniae. Among the 1,307 pigs, the detection rates of S. suis, G. parasuis, P. multocida, and B. bronchiseptica were higher in virus-positive pigs, especially G. parasuis and P. multocida were significantly (p < 0.01) higher than in virus-negative pigs. In addition, a strong positive correlation was found between coinfection by PRRSV and G. parasuis (OR = 2.33, 95%CI: 1.12~2.14), PRRSV and P. multocida (OR = 1.55, 95%CI: 1.12~2.14), PCV2 and P. multocida (OR = 2.27, 95%CI: 1.33~3.87), PRRSV-PCV2 and S. suis (OR = 1.83, 95%CI: 1.29~2.60), PRRSV-PCV2 and G. parasuis (OR = 3.39, 95%CI: 2.42~4.74), and PRRSV-PCV2 and P. multocida (OR = 2.09, 95%CI: 1.46~3.00). In summary, PRRSV, PCV2, S. suis, and G. parasuis were the major pathogens in pigs with PRDC, and coinfections of two or more PRDC-related pathogens with strong positive correlations were common in China, such as PRRSV and G. parasuis, PRRSV and P. multocida, PCV2 and P. multocida, and also PRRSV-PCV2 and G. parasuis and PRRSV-PCV2 and P. multocida.

ICEHpsaHPS7, a Novel Multiple Drug Resistance Integrative Conjugative Element in Glaesserella parasuis.

Integrative conjugative elements (ICEs) are a kind of novel self-transmissible mobile genetic element. In this study, a novel ICE was identified in Glaesserella (Haemophilus) parasuis We confirmed that it could mediate the migration of antimicrobial resistance genes in G. parasuis and found that there may have been a transferring potential between different serovar strains of G. parasuis These findings demonstrate that the ICE is crucial to the horizontal transfer of antimicrobial resistance among G. parasuis strains.

Pigs Overexpressing Porcine ß-Defensin 2 Display Increased Resilience to Glaesserella parasuis Infection.

As the causative agent of Glässer's disease, Glaesserella (Haemophilus) parasuis has led to serious economic losses to the swine industry worldwide. Due to the low cross-protection of vaccines and increasing antimicrobial resistance of G. parasuis, it is important to develop alternative approaches to prevent G. parasuis infection. Defensins are host defense peptides that have been suggested to be promising substitutes for antibiotics in animal production, while porcine ß-defensin 2 (PBD-2) is a potent antimicrobial peptide discovered in pigs. Our previous study generated transgenic (TG) pigs overexpressing PBD-2, which displayed enhanced resistance to Actinobacillus pleuropneumoniae. In this study, the antibacterial activities of PBD-2 against G. parasuis are determined in vitro and in the TG pig model. The concentration-dependent bactericidal activity of synthetic PBD-2 against G. parasuis was measured by bacterial counting. Moreover, after being infected with G. parasuis via a cohabitation challenge model, TG pigs overexpressing PBD-2 displayed significantly milder clinical signs and less severe gross pathological changes than their wild-type (WT) littermates. The TG pigs also exhibited alleviated lung and brain lesions, while bacterial loads in the lung and brain tissues of the TG pigs were significantly lower than those of the WT pigs. Additionally, lung and brain homogenates from TG pigs possessed enhanced antibacterial activity against G. parasuis when compared with those from the WT pigs. Altogether, these proved that overexpression of PBD-2 could also endow pigs with increased resilience to G. parasuis infection, which further confirmed the potential of using the PBD-2 coding gene to develop disease-resistant pigs and provided a novel strategy to combat G. parasuis as well.

Update on Glässer's disease: How to control the disease under restrictive use of antimicrobials.

Antimicrobials have been commonly used to control bacterial diseases in farm animals. The efficacy of these drugs deterred the development of other control measures, such as vaccines, which are currently getting more attention due to the increased concern about antimicrobial resistance. Glässer's disease is caused by Glaesserella (Haemophilus) parasuis and affects pork production around the world. Balance between colonization and immunity seems to be essential in disease control. Reduction in antimicrobial use in veterinary medicine requires the implementation of preventive measures, based on alternative tools such as vaccination and other strategies to guarantee a beneficial microbial colonization of the animals. The present review summarizes and discusses the current knowledge on diagnosis and control of Glässer's disease, including prospects on alternatives to antimicrobials.