http://www.geocities.com/CapeCanaveral/3504/gallery.htm

Treponema pallidum (syphilis)

SPIROCHAETE BACTERIA

LEPTOSPIROSIS• "rice-field fever" and "cane-cutters

fever" and "swine herder's disease“.

• SYMPTOMS: high fever, severe headache, chills, muscle aches, vomiting, jaundice (yellow skin and eyes), red eyes, abdominal pain, diarrhea, or a rash.

• If the disease is not treated, the patient could develop kidney damage, meningitis (inflammation of the membrane around the brain and spinal cord), liver failure, and respiratory distress. In rare cases death occurs.

http://www.vetmed.wisc.edu/pbs/zoonoses/Leptospira/leptoindex.html

http://nature.org/wherewework/northamerica/states/california/explore/

LEPTOSPIROSIS RESERVOIRS

Specific serovars are "hosted-adapted" to particular reservoir species and generally do not cause disease in those hosts, e.g.:

- L. canicola/dogs- L. icterohaemorrhagiae/rats

- L. grippotyphosa/voles, raccoons and other small mammals- L. bratislava/pigs, and rats and other small mammals

Leptospires can persist in the renal tubules without causing disease, and can be excreted in the urine for very prolonged

periods of time.

Rats are the most common source of infection for humans worldwide.

In the U.S., however, the most significant sources of infection for humans are dogs > livestock > rodents > wild

mammals.

NO VECTORS INVOLVED IN LIFE CYCLE!

http://www.vetmed.wisc.edu/pbs/zoonoses/Leptospira/leptoindex.html

http://www.leptospirosis.org/bacteria/

BORELLIA: LYME DISEASE• SYMPTOMS: "bull's-eye" rash,

erythema migrans, accompanied by nonspecific symptoms such as fever, malaise, fatigue, headache, muscle aches (myalgia), and joint aches (arthralgia).

The incubation period from infection to onset of erythema migrans is typically 7 to 14 days but may be as short as 3 days and as long as 30 days.

http://www.cdc.gov/ncidod/dvbid/lyme/

http://www.cdc.gov/ncidod/dvbid/lyme/

VECTOR LIFE CYCLE:

Are Lizards Lethal?

• Lizards, rodents, and birds are all hosts for Ixodes ticks in Northern California.

• Borrelia burgdorferi infection is 3-4 times greater in nymphal ticks than adult ticks.

• Borrelia are rarely identified in lizard blood… are lizards lethal for Borrelia?

Eisen, Eisen, and Lane, 2004. The roles of birds, lizards, and rodents as hosts for the western black-legged tick Ixodes pacificus. Journal of Vector Ecology 29(2): 295-308.

Lane and Quinstad, 1998. Borreliacidal factor in the blood of the western fence lizard (Sceloporus occidentalis). Journal of Parasitology 84(1): 29-34.

Borreliacidal Factors in Lizard Blood!

• 10 lizards were exposed to 10 nymphs having 78% Borrelia prevalence.

• After feeding and molting, the none of the adult ticks contained Borrelia!

• In contrast, 55% of infected nymphs that fed on rabbits passed Borrelia to adult ticks.

• Borrelia in lizard blood died within 1 hr as compared to 72 hrs in mouse blood. Boiling the lizard blood inactivated the borreliacidal response and the Borrelia were then able to survive for the 72 hrs!

WHERE DOES LYME DISEASE OCCUR?

Not your everyday bacteria!

-Inner and outer membranes.

-Periplasmic flagella.

-Flexible cell wall.

-No LPS.

-Linear chromosome.

-Many plasmids.

-Complete genome known.

-Difficult to genetically manipulate.

-In vitro different than in vivo?

-Mouse-tick experimental model mimics natural infection exists.

OspA -> OspC expression.

Ticks eat for 3-7 days, and only efficiently transmit Borrelia after 2 days of feeding.

Larvae and nymphs can also be infected by Borrelia exposure via capillary tube, microinjection, or immersion.

LABORATORY MANIPULATION

• IN VIVO: – Culture Borrelia within a dialysis membrane in

a rat abdomen.– Borrelia can also be isolated from tissues of

infected animals in limited amounts.

• IN VITRO:– Culture Borrelia using BSK growth media but

colonies take 1-2 weeks to appear and a transformation experiment may take 1 month.

GENETIC STUDIES

• Selectable markers: antibiotic resistance genes such as gyrB, ermC, kan. Avoid generating ampicillin and tetracycline resistant Borrelia since those antibiotics are used in disease treatment.

• Strains: non-infectious strains utilized most often because they are easier to transform. Tradeoffs?

GENETIC MANIPULATION• Allelic recombination:

– Electroporation with gene inactivation constructs and antibiotic resistance reporter.

• Shuttle vectors:– Broad host range vector for autonomous replication of

introduced DNA.– Stability important.– Green fluorescent protein (GFP) also used as reporter.

• Transposon mutagenesis:– Random tagged mutagenesis in genome possible.– Localized saturated mutagenesis still not possible.

Allelic Exchange

Shuttle Vectors

Transposon mutagenesis

TRANSFORMATION

• Electroporation or chemical methods.

• Low frequency and efficiency.

• Large amounts of DNA required.

• Plasmids may have restriction-modification systems that can prevent transformation.

• Infectious clones are harder to work with than non-infectious clones… so limited conclusions can be drawn as to mutant Borrelia virulence and pathogenicity.

BORRELIA PLASMIDS• Largest number of plasmids in any known

bacterial genome.

• Encode essential functions such as resT telomere resolvase on cp26 plasmid.

• Unstable plasmid loss of essential lp25 in vitro -> reduced infectivity in vivo.

• Stable plasmids have non-essential genes such as chb and OspC…only active under certain conditions?

QUORUM SENSING?

• Borrelia has a luxS homologue.

• Borrelia luxS complemented E.coli luxS deficiency and altered Borrelia protein synthesis…

• But no autoinducers were detected in supernatant of Borrelia cultures… is quorum sensing only active under certain conditions?

It all began in Lyme, Connecticut!