Abstract. Although Rocky Mountain spotted fever was documented in northern Mexico during the 1940s, spotted fever group (SFG) rickettsioses.
Am. J. Trop. Med. Hyg.. 55(2). 1996, pp. 157—159 Copyright C 1996 by The American society of Tropical Medicine and Hygiene
UNRECOGNIZED SPOTTED FEVER RICKETTSIOSIS MASQUERADING AS DENGUE
GROUP FEVER IN MEXICO
JORGE E. ZAVALA-VELAZQUEZ, XUE-JIE YU, @t'iD DAVID H. WALKER Faculty
of Medicine, Universidad Autonoma de Yucatan, Menida, Yucatan, Mexico; Center for Tropical Diseases, University of Texas Medical Branch, Galveston, Texas
Abstract. Although Rocky Mountain spotted fever was documented in northern Mexico during the 1940s, spotted fever group (SFG) rickettsioses have subsequently received little attention in Mexico. In this study, sera collected in 1993 from 50 patients from the Mexican states of Yucatan and Jalisco, who were suspected clinically to have dengue fever but had no antibodies to dengue virus, were examined by indirect immunofluonescence for 1gM antibodies reactive with Rickettsia nickettsii, R. akani, and R. typhi. Twenty (40%) of the patients' sera contained IgM antibodies to SFG rickettsiae at a titer of 128 on greater. Among five sera reactive only against R. akari, four were from patients in Jalisco where a cluster of cases occurred in June and July. Among five sera reactive only with R. rickettsii, all were from Yucatan patients. Sera of 10 patients contained antibodies reactive with antigens shared by R. rickettsii and R. akari. The clinical signs and symptoms (fever, 100%; myalgia, 95%; headache, 85%; rash, 85%) were similar to those of dengue fever patients identified in this study. However, the incidence of rash was substantially higher than the nondengue, nonrickettsiosis patients. One or more SFG rickettsioses appear to be present in areas of Mexico not previously recognized to harbor these organisms. The etiologic agent or agents are as yet unknown. In tropical and subtropical areas, acute, flu-like, febrile illnesses with or without an exanthem may represent endem ic or epidemic
infections
such
as dengue
fever,
other
agnosis
arbo
viral, arenaviral, or enteroviral infections, malaria, typhoid fever, leptospirosis, relapsing fever, or other infectious dis eases. In tropical areas of the Western hemisphere, rickettsial diseases are seldom considered or investigated. In Mexico, studies of typhus and spotted fever group (SFG) rickettsioses received substantial emphasis from the 1930s until the the 1950s. Rocky Mountain spotted fever was
identified
in Sinaloa,
where Rhipicephalus 4 An
isolate
of
SFG
Sonora,
sanguineus rickeusia
was
Coahuila,
and
tistics
established
enzyme
no
antibodies
to
dengue
Package
by Stanton
A. Glantz,
McGraw
strain), R. akani (Kaplan strain), and R. typhi strain) were cultivated in Vero cells. Rickett cells were harvested when 100% of the cells The infected cells were centrifuged at 12,000
x g for 30 mm. The pellet of cells was resuspended in half
Sera. During 1993 in a study of dengue fever in Yucatan, blood samples from 38 patients suspected on the basis of clinical signs and symptoms to have dengue fever were eval uated and found to be negative for antibodies to dengue vi test and 1gM capture
contain
of the original volume of 0.1 M phosphate-buffered saline (PBS, pH 7.4) containing 2% sodium azide. Ten microliters of rickettsiae-infected cells were applied onto each well of 12-well antigen slides.6 The antigens on the slides were air dried and then fixed in acetone for 10 mm. The slides were kept at —20°C until used. Patients' sera were diluted in two fold increments with PBS. Ten microliters of diluted sera were added to each well of the antigen slides. The slides were incubated at 37°Cfor 30 mm in a humidity box. The slides were rinsed once, then washed twice for 10 mm per wash in PBS. The slides were incubated with fluorescein isothiocyanate—labeled goat anti-human 1gM (p.-specific) (Kirkegaard & Perry Laboratories, Inc., Gaithersburg, MD) and washed as described above. The slides were mounted with 90% glycerin in PBS under coverslips. The slides were read using an ultraviolet microscope (Nikon, Inc., Garden City, NY) at 400X magnification.
MATERIALS AND METHODS
hy hemagglutination
Computer
(Sheila Smith (Wilmington siae-infected were infected.
Am
blyomma cajennense ticks in the tropical Gulf Coast state of Veracruz.5 Subsequently, few investigations of nickettsial diseases in Mexico have been reported. By 1947, Rickettsia pnowazekii had been isolated in five states of Mexico as well as in Mexico City.4 Rickeitsia typhi isolates had been established from 12 states (Guerrero, Hi dalgo, Jalisco, Mexico, Michoacan, Nayarit, Nuevo Leon, Oaxaca, Puebla, Queretaro, Tamaulipas, and Zacatecas). There was serologic evidence of typhus group infection in eight other states, including Yucatan and Jalisco. The purpose of this project was to determine whether spotted fever and typhus group rickettsioses were the cause of acute febrile illness in two different areas of Mexico, the states of Yucatan and Jalisco. Convalescent sera from pa tients with clinical illnesses consistent with dengue fever who were proven not to have dengue viral infections were examined for evidence of rickettsial infection.
ruses
to
Indirect irnmunofluorescence assays for antihodles to R. rickettsii, R. akari, and R. typhL Rickettsia nickettsii
Durango, from
shown
Hill, Inc.
ticks serve as the vecton.1 also
were
viruses. Clinical data. Information regarding the presence or ab sence of a series of signs and symptoms was obtained from each patient. Proportions of patients with SFG nickettsiosis, dengue fever, and neither diagnosis were compared for sta tistically significant differences in occurrence by the com parison of two properties program in the Pnimer of Biosta
RESULTS
Among the 50 convalescent sena, 20 contained antibodies to SFG rickettsiae at titers ranging from 128 to 2,048 for R. rickettsii and from 128 to 256 for R. akari (Table 1). In five
im
munoassay. An additional 12 sera from dengue-suspect pa tients in Guadalajara, Jalisco submitted for dengue serodi
157
158
ZAVALA-VELAZQUEZ TABLE I
TABLE 2
Demographic data and results of indirect immunofluorescence anti
Percentage of Mexican patients with different signs and symptoms compared according to the serologic evidence of spotted fever group (SFG) rickettsiosis and dengue fever
body tests for antibodies
to rickettsiea
in sera from patients
with
acute febrile illness in Mexico*
Patient
Age (years)
Sex
Date of illness serum col lectedt
1
26
F
17
2
31
F
14
3 4
44 37
F F
10 19
5
46
F
10
6
56
F
15
7
52
M
7
8 9 10 11 12 13 14
NA 22 NA 35 36 25 25
NA M F F F M M
NA 16 3 17 1 4 10
15
26
M
9
16 17 18 19 20
22 14 34 22 46
F F F F M
7 18 13 6 7
SF0rickettsiosis Titer of antibodies Signs and symptoms State of origin
Yucatan Yucatan Yucatan Yucatan Yucatan Yucatan Jalisco Jalisco
Jalisco Jalisco Yucatan Yucatan Yucatan Yucatan Yucatan Jalisco
Yucatan Yucatan Jalisco Yucatan
(n30)100949795949085696733633385*6927*605640705757353133St38t1015 = 20)Dengue (n = 16)Neither (n =
R. rick etlsii
R. akari
R. typhi
2,048
Neg4 Neg. Neg. Neg. Neg. 256 256 128 128 128 256 128
Neg. Neg. Neg. Neg. Neg. Neg. Neg. Neg Neg. Neg. Neg. Neg. Neg. Neg. Neg. Neg. Neg. Neg. Neg. 256
256 128 128 128 Neg. Neg. Neg. Neg. Neg.
1,024 512 512 256
256 256 128 I28 128 512
256 128 128 256
256 256 256
256
a NA = Not available.
t Numberof days after onset of symptoms. @
AND OThERS
Negative at a dilution of I :64.
@ patients, antibodies were detected only against R. akari: four of these patients were from the state of Jalisco. In five pa tients, antibodies were detected against only R. nickettsii; all were from Yucatan. In 10 sera, antibodies were detected against both R. rickettsii and R. akari; one of these sera also had antibodies reactive with R. typhi. In six of these sera, the highest titer was against R. rickettsii; in three, the titer for R. akari was 256 and for R. rickettsii 128. In one patient in whom the antibody titer to R. rickettsii was 5 12, the pres ence of antibodies reactive with R. zyphi was suspected to have resulted from a cross-reaction stimulated by an SFG rickettsia as has been described previously.6 Comparison of the signs and symptoms of 20 patients with SFG rickettsiosis, 30 with neither dengue fever nor rickett siosis, and 16 with dengue fever in this study in Mexico revealed similar clinical manifestations for both diseases (Table 2). However, comparison of the SFG rickettsiosis pa tients with those dengue-suspect patients having neither rick ettsiosis nor dengue revealed a higher proportion of SFG rickettsiosis patients with rash, one of the hallmak manifes tations of Rocky Mountain spotted fever, rickettsialpox, and boutonneuse fever. The patients with dengue fever had a higher incidence of vomiting than the SFG rickettsiosis pa tients. None of the other pairwise comparisons of signs and symptoms differed significantly (P > 0.05). The seasonal distribution of illnesses diagnosed serologi cally as SFG rickettsiosis in Yucatan was throughout the year with only the months of December and January having no cases. In contrast, dengue fever occurs in Yucatan mainly from September through February. The SFG rickettsiosis cases in Jalisco were clustered in June and July of 1993, suggesting an outbreak.
Fever Myalgia Headache Nausea Rash
Chills Eye pain Photophobia Vomiting
Cough Abdominal
pain
Petechiae Epistaxis a The occurrence of rash in SF0 rickeusiosis was significantly greater (P < 0.01) than in nonSFG rickettsiosis, nondengue patients in this study. t The occurrence of vomiting was significantly greater (P = 0.04) in patients with dengue fever than in SF0 rickeusiosis.
DISCUSSION
Rickettsial diseases have received little investigative at tention in tropical areas of the Western hemisphere. Endemic Rocky Mountain spotted fever has been identified in north cnn Mexico,' Costa Rica,7 Panama,8 Colombia,9 and Bra zil'°because its high fatality-to-case ratio has demanded in vestigation. Less severe SFG rickeusioses appear to exist for long periods without discovery even in countries with a high level of scientific expertise in rickettsiology (e.g., R. japon ica in Japan
and R. honei
in
al'
‘12 It is possible
that
SFG rickettsioses with low mortality occur unrecognized in tropical Mexico and elsewhere in Central and South Amen ica. The ability of SFG rickettsiae to be maintained in trop ical locations is demonstrated by R. conorii in Ethiopia, So malia, Kenya, Zimbabwe, and India, R. australis in Queens land, and unidentified SFG rickettsiae causing human infec tion in Thailand, Taiwan, and Hainan Island.'@'9 The organism(s) that stimulated antibodies to SFG rick ettsiae in the 20 patients in this study is presently unknown. The tropical isolate of a SFG rickettsia from A. cajennense ticks in the Veracruz state of Mexico was identified as R. nickettsii
on
the
which is known phali,
R. parkeri,
basis
of
cross-protection
to be conferred and some
other
of
by R. conorii, SFG
rickettsiae
guinea
pigs,
R. rhipice as well
as
by R. rickettsii, but not by R. akari.5 2022 To the best of our knowledge, no human cases of SF0 rickettsiosis or isolates of SFG rickettsiae from humans have been described from Veracruz. Until human isolates are established from Mexican patients in Yucatan and Jalisco, the identity of the pathogen will be problematic. The etiologic agent stimulating the an tibodies to R. rickettsii in patients in Yucatan could conceiv ably be a strain of R. rickettsii of low virulence, as appar ently affected patients in Idaho with a mortality rate of ap proximately 3% in the era of the l890s to the 19205,23 24R. conorii, or another SFG rickettsia, possibly even a novel species. It would not be too surprising if R. conorii-infected ticks had been introduced accidentally into Mexico by Span ish conquistadores on animals brought from Spain. A pro
SFG RICKETTSIOSISIN MEXICO spective clinical study is needed to evaluate febrile exan thematous disease in Mexico for the presence of eschars. It is noteworthy, however, that eschars were not noted during the first decade after boutonneuse fever was described in the Mediteranean region and are seldom observed in SFG rick ettsiosis patients in Israel and Zimbawe currently. 13.25-27 The serologic and epidemiologic differences observed in Jalisco merit separate consideration. The temporal clustering and predominant reactivity with R. akani suggest an outbreak of rickettsialpox. Epidemiologic investigation and prospec tive clinical evaluation for the presence of eschar and vesic ular rash and isolation of rickettsiae are required to deter mine whether rickettsialpox occurs in Mexico. The recent isolation of R. akani from a patient in Croatia illustrates how little is known of the geographic distribution of this neglect ed pathogen.28 The demonstration that 20 of 50 undiagnosed febrile pa tients in Mexico had evidence considered diagnostic of an SF0 rickettsiosis emphasizes that an illness presumably re sponsive to doxycycline treatment is masquerading as den gue fever in the tropical Western hemisphere. Acknowledgments: We thank Dr. Jose A. Farfan A. and Maria Alba Lorono of the Arbovirus Laboratory of the Dr. Hideyo Noguchi Center for Research of the Autonomous University of the Yucatan for scientific contributions, and Josie Ramirez for expert secretarial assistance in the preparation of this manuscript. Financial support: This study was supported by a research grant from the National Institute of Allergy and Infectious Disease (AI-2 1242). Authors' addresses: Jorge E. Zavala-Velazquez, Faculty of Mcdi cine, Universidad
Autonoma
de Yucatan,
Merida,
Yucatan,
Mexico.
Xue-Jie Yu and David H. Walker, Department of Pathology, Center for Tropical
Diseases,
University
of Texas Medical
Branch, 301 Uni
versity Boulevard, Galveston, TX 77555-0609.
study of Rocky Mountain
8. Calero
spotted
MC,
Nunez
JM,
Goytia
RS,
1952.
Rocky
Mountain
spotted fever in Panama. Report of two cases. Am J Trop Med Hyg 1: 63 1—636.
9. Patino L, Afanador A, Paul H, 1937. A spotted fever in Tobia, Columbia. Am J Trop Med 17: 639—653. 10. Dias E, Martins AV, 1939. Spotted fever in Brazil. A summary. Am J Trop Med 19: 103—108.
11. Uchida T, Uchiyama T, Kumano K, Walker DH, 1992. Rickett sia japonica
sp. nov., the ecological
agent of spotted
fever
group rickeusiosis in Japan. Int J Syst Bacteriol 42: 303—305. 12. Baird RW, Lloyd M, Stenos J, Ross BC, Stewart
RS, Dwyer B,
1992. Characterization and comparison of Australian human spotted
fever group rickeusiae.
J Clin Microbiol
30: 2896—
2902. 13. Kelly PJ, Beati L, Matthewman LA, Mason PR, Dasch GA, Raoult D, 1994. A new pathogenic spotted fever group rick ettsia from Africa. J Trop Med Hyg 97: 129—137. 14. Walker DH, Liu Q-H, Yu X-J, Li H, Taylor C, Feng H-M, 1992.
Antigenic diversity of Ricketisia cononii. Am J Trop Med Hyg 47: 78—86.
15. Williams WJ, Radulovic S. Dasch GA, Lindstrom J, Kelly DJ, Oster CN, Walker DH, 1994. Identification of Rickettsia con orii infection by polymerase chain reaction in a soldier re turning from Somalia. Clin Infect Dis 19: 93—99. 16. Andrew R, Bonnin JM, Williams 5, 1946. Tick typhus in North
Queensland. Med J Aust 2: 253—258. 17. Sirisanthana
T, Pinyopornpanit
V, Sirisanthana
V. Strickman
D,
Kelly DJ, Dasch GA, 1994. First cases of spotted fever group rickettsiosis in Thailand. Am J Trop Med Hyg 50: 682—686. 18. Takada N, Fujita H, Yano Y, Huang W-H, Khamboonruang C,
1993. Serosurveys of spotted fever and murine typhus in local residents of Taiwan and Thailand compared with Japan. Southeast 19. Feng
H-M,
Asian J Trop Med Public Health 24: 354—356. Chen
T-S,
Lin B-H,
Lin Z-Y, Wang
P-F, Su Q-H,
Xia H-R, Kumano K, Uchida T, 1991. Serologic survey of spotted fever group rickettsiosis on Hainan Island of China. Microbiol Immunol 35: 687—694. 20. Badger LF, 1933. Rocky Mountain spotted
fever and bouton
Public Health Rep 48: 507—51 1. 21 . Lackman DB, Bell FJ, Stoenner HG, Pickens
EG, 1965.
The
Rocky Mountain spotted fever group of rickettsias. Health
1. Bustamante ME, Varela G, 1943. Una nueva rickeusiosis en Mexico: Existencia de la fiebre manchada americana en los estados de Sinaloa y Sonora. Rev Inst Salub Enferm Trop 4: 189—210. 2. Mariotte CO. Bustamante ME, Varela G, 1944. Hallazgo del Rhipicephalus sanguineus Latreille infectado naturalmente
con fiebre manchada de las Montanas-Rocosas,
en Sonora
Rev Inst Salub Enferm Trop 5: 297—300. ME, Varela G, Mariotte OC, 1946. II.-Estudios
de
fiebre manchada en Mexico. Fiebre manchada en la Laguna. Rev Inst Salub Enferm Trop 7: 39—48. 4. Bustamante ME, Varela G, 1947. Distribucion de las rickett siasis en Mexico. Rev Inst Salub Enferm Trop 8: 3—14. 5. Bustamante ME, Varela G, 1946. III.-Estudios de fiebre man chada en Mexico: Hallazgo del Amblyomma cajennense na
turalmente infectado, en Veracruz. Rev Inst Salub Enferm
Lab Sci 2: 135—141. 22. Gage KL, Jerrells TR, 1992.
Demonstration
and partial
char
acterization of antigens of Rickettsia rhipicephali that induce cross-reactive cellular and humoral immune responses to Rickettsia rickettsii. Infect Immun 60: 5099—5106. 23. Maxey EE, 1899. Some observations of the se-called spotted
fever of Idaho. Med Sentinel 7: 433—438. 24. Parker RR, Spencer RR, 1930. Results of four years' vaccination. Studies on Rocky Mountain Spotted
human Fever.
Washington DC: United States Government Printing Office, U.S. Treasury Department Public Health Service. Hygienic Laboratory
Bulletin
154: 72—103.
25. Conor A, Bruch A, 1910. Une fievre eruptive observee en Thn isie. Bull Soc Pathol
Exot 3: 492—496.
26. Pieri J, 1933. La Fievre Exanthematique dii Littoral Mediter raneen ou Fievre Boutonneuse. Paris: G. Doin & CIE. 27. Shulchynska H, Dagan R, Schlaefer F, Keynan A, 1982. Spotted
fever in the Negev. Harefuah 102: 317—319.
Trop 7: 75—78.
6. Philip RN, Casper EA, Ormsbee RA, Peacock MG, Burgdorfer W, 1976. Microimmunofluorescence test for the serological study of Rocky Mountain Microbiol 3: 5 1—61.
L, 1986. Ecological
fever in Costa Rica. Am J Trop Med Hyg 35: 192—196.
neuse fever. A study of their immunological relationship.
REFERENCES
(Mexico). 3. Bustamante
7. Fuentes
159
spotted
fever and typhus.
J Clin
28. Radulovic
5, Feng
H-M,
Morovic
M, Djelalija
B, Popov
V.
Walker DH, 1996. Isolation of Rickettsia akari from a patient in an endemic region of Mediterranean Infect Dis 22: 216—220.
spotted
fever.
Clin
