A Mouse Model to Study the Pathogenic Transition of Streptococcus pneumoniae Following Viral Co-Infection

Begin with biofilm-grown Streptococcus pneumoniae, an asymptomatic nasopharyngeal colonizer, suspended in a buffer

Intranasally administer the suspension to an unanesthetized mouse, allowing inhalation into the upper respiratory tract.

In the nasopharyngeal epithelium, the bacteria establish a biofilm that facilitates colonization and suppresses antiviral cytokine production from host cells.

After anesthetization, extend the mouse’s tongue to access the trachea.

Administer the influenza A virus into the trachea to infect the lower respiratory tract.

After the mouse recovers, immediately inoculate the virus intranasally to target the upper respiratory tract.

Viral infection dysregulates the host immune system and damages mucosal tissue, triggering bacterial dispersal into the lower respiratory tract.

In the virus-altered lung environment, the bacteria transition into invasive pathogens, inducing pulmonary inflammation and systemic spread.

The infection also suppresses neutrophil function, reducing bacterial clearance.

This co-infection model enables investigation of host-microbial interactions.

Thaw the biofilm grown aliquots on ice and spin at 1,700 g for five minutes. Carefully remove and discard the supernatant without disrupting the pellet. Then, wash the pellet by resuspending it in 1 milliliter of PBS and spin it again. Remove the supernatant and resuspend the pellet in the volume needed to reach the desired concentration.

Inoculate the C57 black six male mouse intranasally with 5 times 10 to the sixth colony-forming units or CFU by pipetting 5 microliters of the diluted inoculum into each naris. After inoculation, hold the mouse firmly, stabilizing the head until the volume is inhaled. Once the virus has thawed, dilute the virus in PBS to the desired concentration.

Place ophthalmic lubricant on the eyes of the mouse prior to anesthesia. Immediately infect the anesthetized mouse with 50 microliters of 20 plaque-forming units, or PFU, of influenza A virus or IAV intratracheally by using blunt tweezers to pull the tongue out of the mouth and pipetting the volume of liquid down the trachea. Following recovery, intranasally inoculate 10 microliters of 200 PFU of IAV using the inoculation method described earlier.