Optimal growth conditions for Borrelia

Author: LymeRICK , Marie Kroun, MD - upd. Nov. 2008
Original URL for this article: http://lymerick.net/Borrelia-growth-optimum.html

PubMed search, with "Borreli*+optim*+growth" as criteria:
http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Search&db=PubMed&term=Borreli*+optim*+growth

Temperature 

Pubmed search:  http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Search&db=PubMed&term=Borreli*+optim*+growth+temperature

NOTE it seems that the most common European strains B. afzelii and B. garinii has both 4-6 degree C HIGHER growth optimum, than the original US strain Borrelia burgdorferi B31, which did grow well also at temperatures 35-37!
... below 20 degree C there is little chance of obtaining positive culture in modified BSK-II medium.

IMPLICATIONs of the current knowledge:
Whenever culture of Borrelia burgdorferi sensu lato is being attempted, one should keep the temperature in the range of 35-37 degree C, in order to have the best possible chance of getting growth of ALL the 3 in EU common Borrelia strains!

It is noteworthy to mention in this context that many patients report that they experience a drop in temperature concurrent with start of a Borrelia flare of about 0.5 to 1 degree C (first day) concurrent with having chills and/or very cold extemities ....
In my project temperature (measured in rectum) could be noticed dropping from 37-37.5 degree C down to the range 35.5 to 36.5 degree C at begin of flare.
Later (about 6-12 hours after start of flare) the patient may feel like getting a fever, yet measure a temperature of "only" 37.7 degree C

This would have been named FEVER (38.7 degree C) if the temperature increase of 1.7 degree C had been added to a previous level of 37.0 before
- but is only named SUBFEBRILE when the same increase of 1.7 degree C is added to a previously lowered temperature of 36.0 degree C!

Question/theory:
Is it possible that Borrelia spirochaetes are able to manipulate its mammal hosts temperature down to a temperature range that suits it better for its optimum growth?

 

Selected references:

Growth temperature ranges of Borrelia burgdorferi sensu lato strains.
Hubalek Z, Halouzka J, Heroldova M. J Med Microbiol 1998 Oct;47(10):929-32. PMID: 9788818

Three strains of Borrelia burgdorferi sensu lato, representing three human pathogenic genomospecies (B31, B. burgdorferi sensu stricto; BR14, B. garinii; BR75, B. afzelii) were grown in BSK-H medium at different temperatures and the spirochaetal cells were counted by dark-field microscopy after 0, 4, 8, 16 and 48 daysApproximate optimum (minimum-maximum) temperatures for the in-vitro growth were found to be:
Strain Mean Range of temperature
Borrelia burgdorferi sensu strictu, B31 33 degrees C 22-39 degrees C
B. afzelii, strain BR75 35 degrees C 20-40 degrees C
B. garinii, BR14 37 degrees C 20-41 degrees C

Conclusion: Maximum, optimum and minimum growth temperatures seem to be important characteristics of B. burgdorferi s.l. strains, with relevance for the symptomatology, epidemiology and epizoology of Lyme borreliosis.

Note the HIGHER mean optimum temperature in the most common "EU" strains, compared to the first found "US" strain B31!


Growth parameters of Borrelia burgdorferi sensu stricto at various temperatures.
Heroldova M, Nemec M, Hubalek Z. Zentralbl Bakteriol 1998 Dec;288(4):451-5. PMID: 9987182

Growth of Borrelia burgdorferi sensu stricto (prototype strain B-31) was studied in Barbour-Stoenner-Kelly BSK-H liquid medium, supplemented with 4.5% rabbit serum and antibiotics (phosphomycin, rifampicin), at various temperatures to early stationary growth phase. The number of cells was determined by darkfield microscopy. In the range of cultivation temperatures of 25 degrees C to 37 degrees C, generation time was between 8.26 and 12.36 h (the shortest one at 33 degrees C), and the specific growth rate between 0.056 and 0.083 h-1 (the highest one at 33 degrees C). The optimum growth temperature for B. burgdorferi was 33 degrees C, although good growth was also observed at 28 degrees C, 30 degrees C, 35 degrees C and 37 degrees C. The strain grew well but slowly at the temperature of 25 degrees C, whereas no growth was observed at 20 degrees C.            


Clinical implications of delayed growth of the Lyme borreliosis spirochete, Borrelia burgdorferi.
MacDonald AB, Berger BW, Schwan TG. Acta Trop 1990 Dec; 48(2): 89-94. PMID: 1980573

Lyme borreliosis, a spirochetal infection caused by Borrelia burgdorferi, may become clinically active after a period of latency in the host. Active cases of Lyme disease may show clinical relapse following antibiotic therapy. The latency and relapse phenomena suggest that the Lyme disease spirochete is capable of survival in the host for prolonged periods of time.
We studied 63 patients with erythema migrans, the pathognomonic cutaneous lesion of Lyme borreliosis, and examined in vitro cultures of biopsies from the active edge of the erythematous patch.
Sixteen biopsies yielded spirochetes after prolonged incubations of up to 10.5 months, suggesting that Borrelia burgdorferi may be very slow to divide in certain situations.
Some patients with Lyme borreliosis may require more than the currently recommended two to three week course of antibiotic therapy to eradicate strains of the spirochete which grow slowly.   

16/63 (25%)  of patients w/ EM, biopsy for Bb positive on culture.   Biopsies were cultures for up to 12 months! Temp. 35 degr. C [appropiate] for 3 weeks, thereafter temperature was lowered to 24 degr. C [suboptimal], see PMID: 9788818, 9987182)
The time to detect motile spirochetes: 76-319 d (2.5-10.5 mo), mean 181d  (6mo).
When biopsy was left in culture medium vs removed after 24h, spirochetes could be seen sooner, 147d vs 208d.

Citation page 93, last paragraph:
"Antibiotics are only able to kill actively dividing Borrelia. If a borrelial cell does not divide at least once during the period of antibiotic therapy, it may persist in the host and produce relapse or recrudescence of disease. Therefore, some cases of Lyme borreliosis may require prolonged periods of antibiotic therapy to influence those cell lines of B. burgdorferi with a very slow cycle of cell division."   

Anaerobic

Enhanced culture of Borrelia garinii and Borrelia afzelii strains on a solid BSK-based medium in anaerobic conditions.
De Martino SJ et al. Res Microbiol. 2006 Oct;157(8):726-9. PMID: 16814991

The growth of 29 human strains from the three main pathogenic species of Borrelia burgdorferi sensu lato on a solid BSK-based medium was compared in two culture atmospheres: 3% CO(2) air and anaerobiosis.
All strains grew under anaerobic conditions, whereas only 13 strains were able to grow in aerobiosis with 3% CO(2) (P<0.001). In the latter condition, 75% of the B. burgdorferi sensu stricto strains grew versus 33% of the B. garinii and B. afzelii strains. These data suggest that, especially for B. garinii and B. afzelii species, anaerobic conditions enhance growth yield and speed of low-passage Borrelia strains.
 

Well, below reference is NOT about a Borrelia burgdorferi sensu lato strain, yet was included in this list, because it is another Borrelia that also has a relatively high optimal growth temperature of 34-37 degreee C, supporting above statement about implications of current knowledge about temperature optimum for growth of BORRELIAE:
 
A novel, fast-growing Borrelia sp. isolated from the hard tick Hyalomma aegyptium in Turkey.    
Güner ES, Hashimoto N, Kadosaka T, Imai Y, Masuzawa T. Microbiology. 2003 Sep;149(Pt 9):2539-44.  PMID: 12949178    PDF

A novel, fast-growing spirochaete was isolated from the hard tick Hyalomma aegyptium (family Ixodidae, subfamily Metastriata) using Barbour-Stoenner-Kelly (BSK) II medium. Tick samples were taken during the summer of 2000 from the Istanbul area in northwestern Turkey. Sixty-seven of 153 adults (44%) and 72 of 185 nymphs (39%) were infected with the novel spirochaete, whereas none of the 20 larvae examined were infected. The optimal growth temperature of the spirochaete in BSK II medium was 34-37 degrees C, and it could grow at 39 degrees C. Doubling times at 34 and 37 degrees C were 5.3 and 5.1 h, respectively. Six pure cultures of the spirochaete were obtained and characterized by microscopic observation, sequence analysis of the flagellin gene (flaB), SDS-PAGE and Western blotting. The spirochaete was morphologically similar to those of the genus Borrelia and contained a 41 kDa protein reactive with mAb H9724 specific to the flagellin of a Borrelia species. Polyclonal antibody raised to this spirochaete reacted with several antigen bands, whereas no bands were detected with Borrelia burgdorferi, Borrelia hermsii, Borrelia turicatae and Borrelia parkeri. The flaB sequences of the six isolates showed high similarity, with sequence similarity values ranging from 99.2 to 100%; however, the similarity of the isolates' flaB sequences to those of the Lyme-disease-related Borrelia and relapsing-fever-associated Borrelia species was less than 90%. These findings suggest that the unique spirochaete is a member of the genus Borrelia, and differs from previously described Borrelia species.

The optimal growth temperature of this Borrelia spirochaete in BSK II medium was 34–37 degree C, and it could grow at 39 degree C.
 



Questions:
Which is/are the common mammal host for blood meals for this particular Ixodes tick, Hyalomma aegyptium?
Can this new strain cause Lyme-like infectious disease in mammals? - and in that case, which symptoms are caused by it?