2018 Undergraduate Microbiology research poster session

Nine undergraduate students presented in the 2018 Undergraduate Microbiology Research poster session on April 20, 2017, they include the following:

Emily Hoeper (Yahr lab)
Isabella Morbidelli/Robert Nordell IV (dual presentation) (Yahr lab)
Breanna Kramer-Riesberg (Klingelhutz lab)
Thomas Osborne (Salgado-Pabon lab)
Aissa Kergna (Butler lab)
Carly Twigg (Roller lab)2018 UG Research poster session crowd
Yuxun Zhang (Yahr lab)
Sheng Hao Lu (Maury lab)

Congratulations to undergraduate students Emily Hoeper (2017-2018 Stinski Fellow), and copresenters Isabella Morbidelli/Robert Nordell IV, both won the Allen J. Markovetz Award for best poster presentation at this year's undergraduate poster session.


Read the following selected research descriptions from the participants.

Emily Hoeper (Yahr Lab)

Small, non-coding RNAs that modulate Type III Secretion System Gene Expression 
Isabella Morbidelli, Robert Nordell, Emily Hoeper, Yuxun Zhang, Matthew Ferry, Zachary Slusher, Trey Vanhemert, Deven Voelker, Emily Williams McMackin, Kayley Janssen, Timothy Yahr 

Pseudomonas aeruginosa is a Gram-negative bacteria known for causing infection in immunocompromised individuals. A prominent aspect of P. aeruginosa virulence is a type III secretion system (T3SS). T3SSs allow Gram-negative bacteria to Emily Hoepertranslocate effector proteins into the host cell cytoplasm through a needle-like secretion apparatus. This research is focused on determining if T3SS gene expression is modulated by small regulatory RNAs (sRNAs). sRNAs are small non-coding transcripts that post-transcriptionally affect gene expression, often with the aid of Hfq. Hfq is an RNA chaperone that facilitates imperfect basepairing between the sRNA and its target mRNA. The central hypothesis is that sRNAs downregulate and upregulate type III gene expression. Using a transcriptional reporter for T3SS gene expression, we screened a library of sRNA overexpression constructs. Bioinformatics suggest that P. aeruginosa has 513 sRNAs. Our library consisted of 247 sRNAs, selected for being under a 200 nucleotide cutoff and derived from a defined strand of the coding sequence. Out of the 247 sRNAs screened in strain PAK, 5 sRNAs demonstrated a 50% decrease in T3SS activity when compared to a strain carrying a vector control. To verify that these sRNAs have an effect in other P. aeruginosa strains, they were also tested in strains PA103 and PA14. Each of the sRNAs was also tested for activity in an hfq mutant and two distinct classes were identified, Hfq-dependent and -independent. Future studies will examine the molecular basis for regulation by the sRNAs.


Isabella Morbidelli/Robert Nordell IV (dual presentation) (Yahr lab)

Regulation of Pseudomonas aeruginosa Virulence Functions by Small Non-coding RNAs
Isabella Morbidelli*, Robert Nordell*, Emily Hoeper, Yuxun Zhang, Matthew Ferry, Zachary Slusher, Trey Vanhemert, Deven Voelker, Emily Williams McMackin, Kayley Janssen, Timothy Yahr *equal contributions, shared presentation

Abstract: Pseudomonas aeruginosa is an opportunistic nosocomial pathogen known for its virulence in immunocompromised humans. Although P. aeruginosa has 513 predicted sRNAs, few have known functions. The primary goal of this project was to idIsabella Morbidelli/Robert Nordell IVentify sRNAs that regulate P. aeruginosa virulence functions. We specifically sought to identify sRNAs that regulate motility or confer antimicrobial resistance properties. Individual sRNAs were cloned into an expression vector and transformed into strain PAK. Swimming and twitching motility assays were used to identify sRNAs that regulate flagella and type 4 pili function. Measurements for individual sRNAs were compared to strains carrying an empty vector control. Small RNAs that exhibited deviations of more than two-fold from WT strain PAK were assayed in strains PA103, PAO1, and PA14 to confirm the phenotypes. Seven sRNAs had significant deviations in flagella expression while one sRNA had deviations in both flagella and pili expression. Further testing confirmed that three (64, 179, and 206) of the sRNAs had similar effects in all strains. We took a different approach to assess the role of sRNAs in antimicrobial resistance. Hfq is an RNA chaperone that facilitates basepairing between sRNAs and mRNA targets. We hypothesized that screening an Δhfq mutant for changes in antimicrobial resistance might reveal a role for sRNAs. Using a panel of antibiotics we determined the minimum inhibitory concentrations (MICs) for WT strain PA103 and the Δhfq mutant. The Δhfq mutant has a slight growth phenotype and demonstrated increased susceptibility to each of the tested antibiotics. It is thus unclear whether the increased susceptibility represents an inherent property of the Δhfq mutant or a specific defect in sRNA function. In conclusion, our project identified three sRNAs that control motility. The future goal will be to determine the mRNA targets of those sRNAs and the molecular basis for regulation.


Carly Twigg (Roller Lab)

Regulation of Membrane Budding During HSV Nuclear Egress  
Carly Twigg and Richard Roller, Department of Microbiology, University of Iowa

Abstract: Herpesviruses replicate, express and package their genomes in the cell nucleus. They must move DNA-containing capsids from the nucleus to the cytoplasm to complete assembly and then exit the cell. They do this by a process called nuclear egress in which capsids become enveloped at the inner nuclear membrane and then de-enveloped at the outer nuclear membrane. This process is accomplished by a complex of two conserved proteins pUL34 and pUL31. Mutation of a charge cluster in pUL34, CL13, causes a severe growth defect that is correlated with promiscuous budding of the inner nuclear membrane. The CL13 growth phenotype can be suppressed by mutations outside of the pUL31 and pUL34 genes. Two such independent mutants were found to be different in their growth and spread phenotypes. The two mutants have both common and unique single nucleotide polymorphisms (SNPs). The single common SNP is a frameshift in UL35 and a virus was made containing that mutation in addition to CL13 expression. One mutant, Sup7, has a rearrangement in the US region of the genome. The other mutant, Sup9, has a SNP in the intergenic region between US8 and US9.

Yuxun Zhang (Yahr lab)

sRNAs Associated with Motility and Type Three Secretion System in Pseudomonas aeruginosa.Yuxun Zhang

The Gram-negative bacterium Pseudomonas aeruginosa is an opportunistic pathogen normally found in soil and moist environments. Due to the continuing emergence and spread of antimicrobial resistant strains, therapeutic options are increasingly limited. For that reason P. aeruginosa infections are associated with high morbidity and mortality. The type three secretion system (T3SS) is a needle-like complex used to inject toxins into host cells. The T3SS is a critical virulence factor and is associated with an increased mortality among infected patients. Type IV pili and flagella are required for twitching and swimming motility, respectively, and are also required for virulence. To better understand mechanisms that control virulence gene expression we overexpressed a library of 248 small regulatory RNAs (sRNAs) and screened for phenotypic changes in expression of type three secretion system and motility. Several sRNAs that regulate T3SS gene expression and bacterial motility were identified, shown by a significant reduction in swimming and twitching tests, and reporter gene expression. Future studies will examine how the sRNAs mediate regulatory control. Understanding regulation of P. aeruginosa by sRNAs may lead to potential methods to combat infections.



Wednesday, May 2, 2018