The patient's clinical picture is consistent with S. aureus meningitis. S. aureus looks purple after a gram stain ("gram-positive"), because the cell wall retains the violet dye.
The vast majority of bacteria can be divided into gram-positive bacteria and gram-negative bacteria. These terms refer to the color of the bacteria after performing a gram stain, which involves first processing a sample slide with crystal violet dye, and then with a counterstain (often fuchsin). This stain differentiates between bacteria that have a single, membrane bilayer encased with a thick layer of peptidoglycan ("gram-positive"), from bacteria that have two membrane bilayers with a thin layer of peptidoglycan in between ("gram-negative"). This difference is perhaps most clinically relevant when considering antibiotic coverage of different types of organisms. Some antibiotics are not able to penetrate the membrane bilayer of gram-negative bacteria, while other antibiotics are not able to penetrate the thick peptidoglycan layer of gram-positive bacteria.
Seehusen et al. discuss the analysis of cerebrospinal fluid. They note that the sensitivity of Gram-staining is dependent on the causal organism, with nearly 90% sensitivity for S. aureus and less than 50% sensitivity for Listeria. Having a higher number of colony forming units increases the chance of finding bacteria on a gram stain. In addition, nearly 10% of Gram stains are misread, and therefore technician reliability should be considered when accounting for a Gram stain.
Nigrovic et al. perform a retrospective cohort study to evaluate the validity of the Bacterial Meningitis Score for determining whether children with a cerebrospinal fluid pleocytosis actually have bacterial meningitis. The prediction tool defines those at very low risk for bacterial meningitis as those who have none of the following: "positive CSF Gram stain, CSF absolute neutrophil count (ANC) of at least 1,000 cells/microL, CSF protein of at least 80 mg/dL, peripheral blood ANC of at least 10,000 cells/microL, and a history of seizure before or at the time of presentation." They found that children with a pleocytosis who lack all of the above characteristics have a 0.1% chance of having bacterial meningitis.
Figure A shows a Gram stain of S. aureus. Illustration A shows a Gram stain of S. aureus (purple, gram-positive) and E. coli (pink, gram-negative). Illustration B shows the structure of the bacterial cell wall in gram-positive bacteria (A) and gram-negative bacteria (B).
Answers 2-5: These bacterial organelles do not affect Gram staining.
Seehusen DA, Reeves MM, Fomin DA. Cerebrospinal fluid analysis. Am Fam Physician. 2003 Sep 15;68(6):1103-8.
PMID:14524396 (Link to Abstract)
Nigrovic LE, Kuppermann N, Macias CG, Cannavino CR, Moro-Sutherland DM, Schremmer RD, Schwab SH, Agrawal D, Mansour KM, Bennett JE, Katsogridakis YL, Mohseni MM, Bulloch B, Steele DW, Kaplan RL, Herman MI, Bandyopadhyay S, Dayan P, Truong UT, Wang VJ, Bonsu BK, Chapman JL, Kanegaye JT, Malley R; Pediatric Emergency Medicine Collaborative Research Committee of the American Academy of Pediatrics. Clinical prediction rule for identifying children with cerebrospinal fluid pleocytosis at very low ris
PMID:17200475 (Link to Abstract)