Interaction of Mycobacterium avium subsp. paratuberculosis with bovine sperm.

Mycobacterium avium subsp. paratuberculosis (MAP) is responsible for Paratuberculosis mainly affecting domestic ruminants. The interaction between MAP and sperm and/or germ cells has not yet been established, however the adherence between MAP and the host cell surface is associated to the 85 complex proteins that bind to the host cell's fibronectin. Therefore, this study aimed to evaluate the binding of MAP to bovine sperm and to verify changes in these cells by the presence of MAP before and after sperm cryopreservation. Polyclonal antibodies to MAP 85 complex proteins were produced and utilized in the analyzes. Two Nelore bulls were used for semen collection and MAP dilutions (103-108 CFU/mL) were inoculated in the samples; sperm motility and vigor were evaluated using light microscopy at different times before and after cryopreservation and in the presence and absence of the antibodies 85A and 85B. Interaction of MAP and sperm, interaction of MAP and sperm in the presence of Ab 85A and in the presence of Ab 85B were analyzed by scanning electron microscopy. The viability of MAP after sperm cryopreservation were evaluated by plating the samples after thawing. It was observed that sperm in the presence of MAP shows a decrease in motility and vigor, and that the higher the MAP concentration, the lower the sperm performance. It was possible to determine the viability of MAP after cryopreservation in samples of higher concentrations, which demonstrates the potential of transmission of this pathogen through artificial insemination. The interaction of MAP with bovine sperm occurs mainly in the midpiece and may be linked to the proteins 85A and 85B present in the MAP membrane.

Differences in the coinfective process of Staphylococcus aureus and Streptococcus agalactiae in bovine mammary epithelial cells infected by Mycobacterium avium subsp. paratuberculosis.

The interactions between Mycobacterium avium subsp. paratuberculosis (MAP) and the causative agents of bovine mastitis are still relatively unknown. Still, it is suspected that they may contribute to the worsening and persistence of mastitis within the mammary epithelial cells. Considering the growing economic implications of paratuberculosis and subclinical mastitis in dairy herds, this study aimed to determine the coinfection interaction between MAP and S. aureus or S. agalactiae in bovine mammary epithelial cells (MAC-T) in an ex-vivo model. For this purpose, internalisation tests of MAP + S. aureus or MAP + S. agalactiae were performed in MAC-T cells for 10, 30 and 120 min. The qPCR was performed to quantify internalised MAP at the time of exposure. Colony-forming units were counted on BHI agar medium for internalised subclinical mastitis bacteria at each time of infection. Viability tests of MAC-T cells, using the lactate dehydrogenase assay, were performed. The results showed that in the MAC-T cells previously infected by MAP and subsequently by S. aureus, there was a rapid internalisation in the first 10 min, maintaining a higher number of internalised bacteria during all exposure times. Regarding MAP + S. agalactiae, there were no changes in the internalisation patterns. The amount of MAP remained constant at all times evaluated, and there was no compromise in the viability of MAC-T cells during the tests. Thus, the results demonstrate the existence of an interaction between MAP + S. aureus, favouring internalisation and being able to contribute to the persistence of subclinical mastitis in dairy herds.

Antimicrobial and Synergistic Activity of 2,2',4-Trihydroxybenzophenone Against Bacterial Pathogens of Poultry.

In poultry farming, the spread of bacterial pathogens results in disease outbreaks causing significant economic losses to this industry. Many of these pathogenic bacteria are zoonotic and have a substantial impact on public health. Antimicrobials are essential for the prevention and treatment of these bacterial infections. However, the indiscriminate use of these agents provides favorable conditions for selection, propagation and persistence of bacteria and development of antimicrobial resistance. We developed a new antimicrobial candidate that could be used alone or in synergy with research protocols for therapeutic, prophylactic and growth promoter uses in the poultry industry. The present study aimed at evaluating the antimicrobial activity of the synthetic compound 2,2',4-trihydroxybenzophenone against pathogenic bacteria that cause important diseases in poultry and public health. We tested the hemolytic effect of this compound, studied its synergistic effect with conventional antimicrobials and analyzed the site of action on the bacteria. The results of our study showed antimicrobial activity of benzophenone against Gram-positive and Gram-negative bacteria with a similar effect in ATCC (American type culture collection) and field isolates. This compound was non-hemolytic. 2,2',4-trihydroxybenzophenone acted on the bacterial cell wall. We identified the synergistic effect between 2,2',4-trihydroxybenzophenone and bacitracin, this effect indicate that antimicrobial synergism may be useful for the treatment of necrotic enteritis in poultry. This compound may also be used as a growth promoter by reducing the dose of bacitracin and thus decreasing the pressure of bacterial resistance in poultry which would circumvent the development of cross-resistance in humans.