Antibiotic resistance in Neisseria gonorrhoeae Karen Cloud-Hansen, PhD March 12, 2007

What is gonorrhea?  Clap, Drip  Sexually transmitted infection caused by the bacterium Neisseria gonorrhoeae  Can be detected by microscopy, culturing, or DNA amplification

Who gets gonorrhea?  Spread by sexual contact  Those at highest risk are young adults (women 15 – 19 and men 20 – 24)  Can be passed from mother to newborn

United States Gonorrhea Infection Statistics  Second most- frequently reported STD  ~800,000 people infected annually  Treatment of gonorrhea costs $1 billion/year

What are the symptoms?  Most men who get gonorrhea experience extremely painful urination and pus release from the urethra  Many women do not experience noticeable symptoms

Why is proper detection and treatment of gonorrhea important?  Co-infection can cause increased transmission of other STDs including HIV  If untreated, the bacteria can spread to other sites in the body Pelvic Inflammatory Disease Meningitis Arthritis Blindness

What are some of the difficulties in treating the disease?  Some people do not know that they are infected  People can be infected with N. gonorrheae multiple times  Can be a difficult topic for some to talk about/embarrassment  Some isolates of N. gonorrhoeae are resistant to antibiotics commonly used to treat gonorrhea

Why not just use other drugs to treat gonorrhea?  Treatment with ciprofloxacin is Inexpensive Only 1 dose needed Medication is delivered orally  Few other antibiotics available

A History of Antibiotic Resistance in N. gonorrhoeae  1976 – Penicillinase-producing N. gonorrhoeae described in case reports  1985 – Tetracycline-resistant N. gonorrhoeae described in case reports  1991 – Ciprofloxacin-resistant N. gonorrhoeae first detected by the Centers for Disease Control

General Mechanisms of Antibiotic Resistance  Deactivation of drug by enzymatic cleavage  Less drug enters the bacterium  Concentration of drug within the bacterium is lowered  Target of drug is altered

Mechanisms of Ciprofloxacin Resistance in N. gonorrhoeae  Deactivation of drug by enzymatic cleavage  Less drug enters the bacterium Changes to Porins  Concentration of drug within the bacterium is lowered Multi-drug efflux pumps  Target of drug is altered

What is the target of Ciprofloxacin?  DNA gyrase (removes supercoils)  Topoisomerase IV (separates daughter strands)  Ciprofloxacin binds to the DNA/enzyme complex forming a physical barrier that prevents movement of the replication fork, RNA polymerase, and DNA helicase

Question  At what stage(s) in gene expression does ciprofloxacin act?  What do you think is the result of treating bacteria with ciprofloxacin?  Write your answers on a sheet of paper

Answers  Ciprofloxacin blocks DNA synthesis. Downstream steps of gene expression do not occur.  Treatment of ciprofloxacin-sensitive bacteria with the antibiotic results in cell death.

What mutations result in increased resistance to ciprofloxacin?  Mutations to near the active site (where the enzyme binds DNA) in DNA gyrase and topoisomerase IV.  Typically, mutations occur in genes encoding one of two parts of each enzyme (gyrA for DNA gyrase and parC for topoisomerase IV).

Genotype Can Perturb Phenotype The following statements describe how a change in genotype can perturb phenotype. Number them in the best sequential order (1=earliest event, 4=latest event): __ Mutant gyrA RNA is translated. __ The gyrA gene of Neisseria gonorrhoeae acquires a mutation because of a replication error. __ Mutant gyrase enzyme is altered, allowing for DNA synthesis to occur even in the presence of ciprofloxacin. __ Mutant gyrA DNA is transcribed.

Genotype Can Perturb Phenotype The following statements describe how a change in genotype can perturb phenotype. Number them in the best sequential order (1=earliest event, 4=latest event): _3_ Mutant gyrA RNA is translated. _1_ The gyrA gene of Neisseria gonorrhoeae acquires a mutation because of a replication error. _4_ Mutant gyrase enzyme is altered, allowing for DNA synthesis to occur even in the presence of ciprofloxacin. _2_ Mutant gyrA DNA is transcribed.

How Do the Centers for Disease Control Monitor Antibiotic Resistance in Neisseria gonorrhoeae? In the United States, much of the information about the emergence of antibiotic resistance in Neisseria gonorrhoeae comes from the Gonococcal Isolate Surveillance Project (GISP). Neisseria gonorrhoeae is isolated from the first 25 men with urethral gonorrhea attending public health clinics in 28 cities. The samples of bacteria are sent to regional laboratories where they are tested for susceptibility to several antibiotics. What are the limits and biases of this study?

Location of GISP Collection Sites STD Clinics Clinics and Regional Labs Birmingham Phoenix Long Beach Los Angeles Orange Co. San Diego San Francisco Denver Miami Atlanta Honolulu Tripler AMC Chicago New Orleans Baltimore Detroit Minneapolis St. Louis Las Vegas Albuquerque Greensboro Cincinnati Cleveland Oklahoma City Portland Philadelphia Dallas Seattle Adapted from CDC slides

Possible Answers  No women are included in the study. There is no way of knowing if antibiotic resistance patterns are the same in Neisseria gonorrhoeae isolated from men and women.  The isolates are taken only from the urethra. There is no way of knowing if the antibiotic resistance patterns are the same in Neisseria gonorrhoeae isolated from the rectum, throat, or other sites in men.

Possible Answers  Epidemiological data would be biased. The patients in public health clinics are often the urban poor. One clinic is located on a military base. This may inflate incidence estimates in certain groups.  The study design misses certain populations including those in rural areas or those seeking treatment from doctors in private practice.