Acute BVDV-2 infection in beef calves delays humoral responses to a non-infectious antigen challenge.

Immunosuppressive effects of an intranasal challenge with non-cytopathic bovine viral diarrhea virus (BVDV) 2a (strain 1373) were assessed through acquired and innate immune system responses to ovalbumin (OVA). Concurrent BVDV infection was hypothesized to delay and reduce the humoral response to ovalbumin (administered on days 3 and 15 post-inoculation). Infected animals followed the expected clinical course. BVDV titers, and anti-BVDV antibodies confirmed the course of infection and were not affected by the administration of OVA. Both the T-helper (CD4(+)) and B-cell (CD20(+)) compartments were significantly (P < 0.05) reduced in infected animals, while the gamma-delta T-cell population (Workshop cluster 1+, WC1(+)) decreased slightly in numbers. Infection with BVDV delayed the increase in OVA IgG by approximately 3 d from day 12 through day 21 post-inoculation. Between days 25 and 37 post-inoculation following BVDV infection the IgM concentration in the BVDV- group decreased while the OVA IgM titer still was rising in the BVDV+ animals. Thus, active BVDV infection delays IgM and IgG responses to a novel, non-infectious antigen.

Une infection aiguë par le BVDV-2 chez les veaux retarde les réponses humorales face à un test à l'aide d'un antigène non infectieux. Les effets immunosuppressifs d'une inoculation défin intranasale à l'aide du virus non cytopathogène de la diarrhée virale bovine (VBVD) 2a (souche 1373) ont été évalués par les réactions acquises et innées du système immunitaire à l'ovalbumine (OVA). On a émis l'hypothèse que l'infection concomitante par le VBVD retardait et réduisait la réaction humorale à l'ovalbumine (administrée aux jours 3 et 15 après l'inoculation). Les animaux infectés ont suivi le cheminement clinique prévu. Les titres de BVDV et les anticorps anti-BVDV ont confirmé le déroulement de l'infection et ils n'ont pas été affectés par l'administration d'OVA. Les compartiments des lymphocytes T auxiliaires (CD4+) et des cellules B (CD20+) étaient significativement réduits (P < 0,05) chez les animaux infectés, tandis que la numération de la population de cellules T gamma-delta (WC1+) a diminué légèrement. L'infection par le VBVD a retardé l'augmentation de l'OVA IgG d'environ 3 jours, à compter du jour 12 jusqu'au jour 21 après l'inoculation. Entre les jours 25 et 37 après l'inoculation suivant l'infection par le BVDV, la concentration d'IgM dans le groupe VBVD a diminué tandis que le titre d'OVA IgM augmentait toujours chez les animaux positifs pour le VBVD. Par conséquent, l'infection active par le VBVD retarde les réactions IgM et IgG face à un antigène non infectieux nouveau.(Traduit par Isabelle Vallières).

Extracellular DNA-induced antimicrobial peptide resistance in Salmonella enterica serovar Typhimurium.

BACKGROUND: The Salmonella enterica serovar Typhimurium PhoPQ two component system (TCS) is activated by low Mg2+ levels, low pH and by antimicrobial peptides (AP). Under Mg2+ limitation, the PhoPQ system induces pmrD expression, which post-translationally activates the PmrAB TCS. PhoPQ and PmrAB control many genes required for intracellular survival and pathogenesis. These include the polymyxin resistance (pmr) operon, which is required for aminoarabinose modification of LPS and protecting the outer membrane from antimicrobial peptide disruption and killing. Extracellular DNA is a ubiquitous polymer in the matrix of biofilms and accumulates in some infection sites. Extracellular DNA chelates cations and thus activates the Pseudomonas aeruginosa PhoPQ/PmrAB systems, leading to expression of the orthologous arn (pmr) operon. RESULTS: Here we show that extracellular DNA induces expression of the S. Typhimurium pmr antimicrobial peptide resistance operon in a PhoPQ and PmrAB-dependent manner. Induction of the pmr genes by DNA was blocked when present with excess Mg2+. Exogenous DNA led to increased resistance of planktonic cultures to aminoglycosides, antimicrobial peptides (AP) and ciprofloxacin, but only AP resistance was PhoPQ/PmrAB-dependent. Extracellular DNA was shown to be a matrix component of S. Typhimurium biofilms cultivated in flow chambers and on glass surfaces. A pmrH-gfp fusion was highly expressed in flow chamber biofilms cultivated in medium with repressing levels of 10 mM Mg2+ and co-localized with eDNA. Expression of pmrH-lux was monitored in plastic peg biofilms and shown to require PhoPQ and PmrAB. Biofilms had higher levels of pmrH expression compared to planktonic cultures. We propose that DNA accumulation in biofilms contributes to the increased pmrH-lux expression in biofilms. CONCLUSIONS: The Salmonella PhoPQ/PmrAB systems and antimicrobial peptide resistance are activated by the cation chelating properties of extracellular DNA. DNA-induced AP resistance may allow immune evasion and increased survival of S. Typhimurium biofilms formed during extracellular growth stages of an infection or outside the host.

Inhibition of bacterial biofilm formation and swarming motility by a small synthetic cationic peptide.

Biofilms cause up to 80% of infections and are difficult to treat due to their substantial multidrug resistance compared to their planktonic counterparts. Based on the observation that human peptide LL-37 is able to block biofilm formation at concentrations below its MIC, we screened for small peptides with antibiofilm activity and identified novel synthetic cationic peptide 1037 of only 9 amino acids in length. Peptide 1037 had very weak antimicrobial activity, but at 1/30th the MIC the peptide was able to effectively prevent biofilm formation (>50% reduction in cell biomass) by the Gram-negative pathogens Pseudomonas aeruginosa and Burkholderia cenocepacia and Gram-positive Listeria monocytogenes. Using a flow cell system and a widefield fluorescence microscope, 1037 was shown to significantly reduce biofilm formation and lead to cell death in biofilms. Microarray and follow-up studies showed that, in P. aeruginosa, 1037 directly inhibited biofilms by reducing swimming and swarming motilities, stimulating twitching motility, and suppressing the expression of a variety of genes involved in biofilm formation (e.g., PA2204). Comparison of microarray data from cells treated with peptides LL-37 and 1037 enabled the identification of 11 common P. aeruginosa genes that have a role in biofilm formation and are proposed to represent functional targets of these peptides. Peptide 1037 shows promise as a potential therapeutic agent against chronic, recurrent biofilm infections caused by a variety of bacteria.

Induction of humoral immune response in piglets after perinatal or post-weaning immunization against porcine circovirus type-2 or keyhole limpet hemocyanin.

The objective of this study was to test the hypothesis that porcine circovirus type-2 (PCV2) vaccination is efficacious when administered in the first week of life. Three groups of pigs were vaccinated with Circumvent either early (at the end of week 1), late (at the end of week 4), or not at all. All 3 groups were later challenged intranasally with PCV2 (at the end of week 5). Two other groups were immunized with keyhole limpet hemocyanin (KLH) as a novel antigen at the end of either week 1 or week 4. Weight, PCV2 genome copy number in serum and saliva, anti-KLH antibody titer, and serum PCV2-neutralizing antibodies were measured weekly. Early PCV2 vaccination or KLH antigen exposure resulted in earlier humoral responses that were slower to develop than in older piglets, yet converged with the responses to later vaccination within 5 wk. Both groups of vaccinated piglets had periods of higher PCV2-neutralizing antibody titers and lower viral levels shortly after weaning and PCV2 challenge, thus supporting the recent labelling of 1 Canadian PCV2 vaccine for use in week 1 and suggesting that early PCV2 vaccination can reduce piglet handling without compromising vaccine efficacy.

L'objectif de la présente étude était de vérifier l'hypothèse que la vaccination contre le circovirus porcin de type 2 (CVP2) est efficace lorsqu'administrée durant la première semaine de vie. Trois groupes de porcs ont été vaccinés avec Circumvent soit hâtivement (à la fin de la semaine 1), tardivement (à la fin de la semaine 4), ou pas du tout. Les trois groupes ont plus tard été inoculés par voie intranasale avec CVP2 (à la fin de la semaine 5). Deux autres groupes ont été immunisés avec de l'hémocyanine de patelle (KLH) à titre de nouvel antigène à la fin de soit la semaine 1 ou la semaine 4. Le poids, le nombre de copies du génome de CVP2 dans le sérum et la salive, le titre d'anticorps anti-KLH, et le titre d'anticorps sériques neutralisants CVP2 ont été mesurés à chaque semaine. La vaccination tôt contre CVP2 ou l'exposition à l'antigène KLH a donné des réponses humorales plus hâtives qui étaient plus lentes à se développer que chez les porcs plus vieux, mais qui convergeaient vers les réponses de la vaccination tardive à l'intérieur d'un délai de 5 sem. Les deux groupes de porcelets vaccinés avaient des périodes de titres d'anticorps neutralisants contre CVP2 plus élevés et des charges virales plus basses peu de temps après le sevrage et le challenge avec CVP2, soutenant ainsi l'étiquetage récent d'un vaccin canadien contre CVP2 pour utilisation dans la semaine 1 et suggérant qu'une vaccination tôt contre CVP2 peut réduire la manipulation des porcelets sans compromettre l'efficacité du vaccin.(Traduit par Docteur Serge Messier).

Calcium chelation by alginate activates the type III secretion system in mucoid Pseudomonas aeruginosa biofilms.

The extracellular biofilm matrix includes primarily DNA and exopolysaccharides (EPS), which function to maintain aggregate structures and to protect biofilms from antibiotics and the immune response. Both polymers are anionic and have cation binding activity, however the impact of this activity on biofilms is not fully understood. Host cell contact is considered the primary signal for activation of most type III secretion systems (T3SS), although calcium limitation is frequently used as a trigger of contact-independent T3SS expression. We hypothesized that alginate, which is a known calcium binding exopolysaccharide produced in mucoid Pseudomonas aeruginosa isolates, can activate the T3SS in biofilms. The addition of exogenous purified alginate to planktonic, non-mucoid PAO1 cultures induced expression of exoS, exoT and exoY-lux reporters of the T3SS in a concentration-dependent manner. Induction by alginate was comparable to induction by the calcium chelator NTA. We extended our analysis of the T3SS in flow chamber-cultivated biofilms, and showed that hyperproduction of alginate in mucA22 mucoid isolates resulted in induction of the exoS-gfp transcriptional reporter compared to non-mucoid paired isolates. We confirmed the transcriptional effects of alginate on the T3SS expression using a FlAsH fluorescence method and showed high levels of the ExoT-Cys(4) protein in mucoid biofilms. Induction of the T3SS could be prevented in planktonic cultures and mucoid biofilms treated with excess calcium, indicating that Ca(2+) chelation by the EPS matrix caused contact-independent induction. However, mucoid isolates generally had reduced exoS-lux expression in comparison to paired, non-mucoid isolates when grown as planktonic cultures and agar colonies. In summary, we have shown a mucoid biofilm-specific induction of the type III secretion system and highlight a difference between planktonic and biofilm cultures in the production of virulence factors.