Neisseria meningitidis is one of the important cause of meningitis and has been extremely susceptible to penicillin. Nevertheless, moderately penicillin resistant strains have been reported in some parts of the world. To our knowledge, there is no such report in Malaysia. We report two clinical isolates that were found to have MICs of decreased susceptibility to penicillin by the agar dilution method.
The indirect immunoperoxidase (HP) test has been used extensively in most government hospitals in Malaysia for the serodiagnosis of scrub typhus, murine typhus and tick typhus during the 1990s. The test was used to determine the IgG and IgM antibody titers in patients' sera for three rickettsial species, ie Orientia tsutsugamushi OT; the causative agent of scrub typhus), Rickettsia typhi (RT; the causative agent of murine typhus), and TT118 spotted fever group rickettsiae (TT; the causative agent of tick typhus). The serological findings obtained from Malaysian hospitals using the IIP test (1994-1999) were analyzed. During the six-year period, a total of 61,501 patients' sera were tested, of which 9.6%, 10.5%, and 12.9% had antibody (IgG and/or IgM of > or = 1:50) for OT, RT and TT respectively. A total of 8.6%, 9.8%, and 9.7% of sera had IgG antibody of > or = 1:50 for OT, RT, and TT respectively, indicating past infection. A total of 3.4%, 3.8%, and 6.4 % of sera had IgM antibody of > or = 1:50 for OT, RT, and TT respectively, indicating recent infection. A total of 2,986 (4.9%), 1,882 (3.1%), and 1,574 (2.6%) of sera had IgG and/or IgM antibody titers of > or = 1:400 for OT, RT, and TT respectively, suggesting active rickettsial infection. The seropositivity rates of OT, RT and TT varied according to geographical locations. While the seropositivity of OT remained constant during the six-year period, a reduction in the seropositivity of both RT and TT was noted during recent years. The serological findings reflect the endemicity of rickettsial diseases, including tick typhus, and endemic typhus in various parts of Malaysia. Awareness of these diseases by health and medical staff and by the general public is important if the mortality and morbidity associated with scrub typhus, tick typhus, and murine typhus in Malaysia, are to be reduced.
In this study, the presence of IgG and IgM antibodies against Borrelia burgdorferi (strain B. afzelii) among Malaysian blood donors and patients admitted to hospital with various infectious diseases was determined. Sera were screened using enzyme-linked immunosorbent assay (ELISA); positive sera were then subjected to Western blot testing. All but one of the blood donors were negative for borrelial antibodies. Of 121 patients' sera, IgM antibodies were detected in 24 (19.8%) and IgG antibodies were detected in 5 (4.1%) sera. Only one of two patients with skin manifestations suggestive of Lyme disease had IgM antibody against B. afzelii. Of 30 patients with exposure to tick typhus, 4 (13.3%) were IgM positive and 1 (3.3%) was IgG positive. Based on the detection of antigenic bands by Western blot, 6 patients' sera showed positive reactions. Antigenic bands of p39, p41 and p59/62 kDa were the commonest findings of Western blotting. This study provides serological evidence of B. afzelii infections in Malaysia; further investigation is needed to correlate serological and clinical findings.
A chromatographic immunoassay cholera antigen detection kit, the Cholera Spot test, was evaluated. The test was found to be specific with a sensitivity of 10(6) cfu/ml for the direct detection of V. cholerae in simulated stool specimens and 10 cfu/ml in simulated cotton-tipped swab specimens after overnight incubation in alkaline peptone water. This enables early recognition of cholera cases and their contacts so that prevention and control measures can be promptly instituted.
The seroprevalence of Orientia tsutsugamushi (OT), Rickettsia typhi (RT) and TT118 spotted fever group rickettsiae (SFGR) among blood donors and febrile Malaysian patients in the urban areas was determined. Of the 240 blood donors, 5.4%, 9.2% and 1.7% had either present or previous exposure to OT, RT and SFG rickettsiae, respectively. Patients admitted to an urban hospital had high seroprevalences of OT (43.5%) and RT (22.9%), as compared to SFGR (11.6%). Antibody levels suggestive of recent infections of scrub typhus, murine typhus and tick typhus were detected in 16.8%, 12.7% and 8.2% of patients respectively. No significant difference was noted in the distribution of rickettsial antibodies among urban patients from 2 geographical locations. However, the serologic patterns of rickettsial infection in the urban areas were different form those of rural areas.
Melioidosis is endemic in Malaysia. Emerging resistance with new and current antimicrobial agents has underscored the need to look further for new antimicrobial agents for the treatment of melioidosis. Hence, we evaluated the in-vitro susceptibility of fifty locally isolated strains of Burkholderia pseudomallei, the causative agent of melioidosis to cefoperazone-sulbactam combination using the method of NCCLS. All the fifty strains tested were susceptible in-vitro to cefoperazone-sulbactam. The MIC90 of the organism for cefoperazone-sulbactam was 4 mg/L. The results of our findings suggested that cefoperazone-sulbactam may be useful in the treatment of melioidosis.
A rapid diagnostic system for scrub typhus using nested polymerase chain reaction (PCR) was applied to clinical samples from Malaysian Aborigines. Whole blood from twenty-four patients suspected of scrub typhus infection were tested using nested polymerase chain reaction and sera were evaluated by the indirect immunoperoxidase test. Antibody responses towards Rickettsia tsutsugamushi were observed in seventeen patients with the majority having high titers of IgG antibodies. Seven patients were seronegative. The nested PCR amplified R. tsutsugamushi DNA from six patients, of which two were negative serologically and four had high titers of IgG antibodies. Second samples collected seven days after treatment were negative by PCR testing. Nested PCR is highly sensitive and specific and may be used to provide rapid confirmation of scrub typhus cases in endemic region.
The diagnostic value of adenosine deaminase (ADA) activity was studied to evaluate its use in the differential diagnosis of tuberculous meningitis in the local setting. Cerebrospinal fluid (CSF) from 119 patients with meningitis and other conditions with central nervous system symptoms were collected and ADA activity determined by the colorimetric method of Guisti read at 628 nm. The CSF was also subjected to other laboratory examinations so as to provide the aetiological diagnosis. All 14 tuberculous meningitis patients had ADA activity greater than the cut off value of 9.0 IU/L. High ADA activity was also seen in 13 of 105 non-tuberculous cases giving a specificity of 87.6%. Even though the ADA activity determination is sensitive for tuberculosis, it was not specific enough to be used as a rapid diagnostic test. However when interpreted together with clinical signs and symptoms and other laboratory tests, it is a useful adjunctive rapid marker for tuberculosis.
The in-vitro susceptibility of quinupristin/dalfopristin, levofloxacin and moxifloxacin against methicillin-resistant Staphylococcus aureus (MRSA) strains, which are also resistant to fusidic acid and rifampicin were carried out to determine whether these antibiotics can be used as an alternative treatment for multiply resistant MRSA strains. The minimum inhibitory concentrations (MIC) of these antibiotics were determined by E-test. Quinupristin/dalfopristin had good activity (MIC90 = 1 mg/L) against these strains while most of the strains showed intermediate resistance to moxifloxacin with MIC90 = 2 mg/L). However, more than 90% of these strains were resistant to levofloxacin with the MICs that ranged from 8 mg/L to 16 mg/L with the majority inhibited at 8 mg/L.
Isolation of rickettsiae from patients' blood samples and organ samples of wild rodents from areas with high seroprevalence of rickettsial infections was attempted using cell culture assay and animal passages. L929 mouse fibroblast cells grown in 24 well tissue culture plate were inoculated with buffy coat of febrile patients and examined for the growth of rickettsiae by Giemsa, Gimenez staining and direct immunofluorescence assay. No rickettsiae were isolated from 48 patients' blood samples. No symptomatic infections were noted in mice or guinea pigs infected with 50 organ samples of wild rodents. There was no rickettsial DNA amplified from these samples using various PCR detection systems for Orientia tsutsugamushi, typhus and spotted fever group rickettsiae.
The pathogenicity of Malaysian isolates of Orientia tsutsugamushi was investigated by a mouse virulence assay. The isolates could be differentiated as low (4 isolates), moderately (3 isolates) and highly virulent (2 isolates) based on the different responses in infected mice. No direct correlation between severity of human scrub typhus infections and virulence of the O. tsutsugamushi in mice was observed. Mice infected with virulent strains of O. tsutsugamushi showed splenomegaly, ascitis accumulation and enlargement of kidneys and livers whereas avirulent O. tsutsugamushi strains were asymptomatic and exhibited ruffled fur for a short period after infection. There was low antibody response in mice infected with isolates of low pathogenicity as compared with those of highly virulent isolates. Upon dissection of the infected mice, enlargement of mouse organs such as spleen, kidney and liver was noted. Presence of rickettsemia in mice was confirmed by the growth of O. tsutsugamushi in the L929 cells when inoculated with blood from infected mice. O. tsutsugamushi was also cultured from the peritoneal exudates of the infected mice. However, DNA of O. tsutsugamushi was only detected in the peritoneal exudates (by PCR) and blood (by cell culture) and not from other tissue samples.
The seroprevalence of various Orientia tsutsugamushi (OT) strains among Malaysian patients with suspected scrub typhus infections was determined using an indirect immunoperoxidase (IIP) assay. IgG against a single OT strain were detected in six sera (3 Karp, 1 Gilliam and 2 TC586), whereas IgM antibodies against a single OT strain (Gilliam) were noted in 3 sera (Gilliam). IgG reactive to all OT strains were present in 33 (47.1%) of the 70 sera and IgM reactive to all OT strains were present in 22 (78.6%) of the 28 sera. The fact that most sera were reactive to multiple OT strains suggests that group-specific antigens are involved in scrub typhus infections, whereas very few were due to strain-specific epitopes present on these strains. Peak IgG and IgM titers were noted more frequently against Gilliam, Karp, and TA763 strains: this suggests that these strains may be the commonest infecting strains among Malaysian patients. Two predominant OT polypeptides consistently reacted with patients' sera were the 70 kDa and 56 kDa proteins.
In this study, recombinant proteins that encompassed the AD I-AD III regions of 56 kDa immunodominant gene of 2 Orientia tsutsugamushi (OT) serotypes; Gilliam and TA763 were expressed in Escherichia coli. Both recombinant proteins exhibited serologic cross-reactivity with the rabbit antisera against various OT serotypes, as evaluated by enzyme-linked immunosorbent assay (ELISA), but not against other rickettsial species, including Rickettsia typhi, R. prowazekii and TT118 SFG rickettsiae. The feasibility of using the recombinant proteins as a diagnostic reagent was further evaluated by ELISA using sera from blood donors and scrub typhus patients. The results suggested a higher affinity of the antihuman IgM than IgG with both recombinant proteins. The IgM ELISA findings were agreeable with the results of indirect immunoperoxidase (IIP) assay especially with sera of high antibody (1:1600). However, more than one antigen are probably needed for development of an effective assay for serodiagnosis of scrub typhus in endemic areas.
Isolation and polymerase chain reaction (PCR) were performed for detection of Mycoplasma pneumoniae from respiratory tract specimens obtained from 200 adult and 200 pediatric patients. M. pneumoniae was isolated from bronchoalveolar lavage fluid of 1(0.5%) adult patient and 4(2.0%) tracheal aspirates of pediatric patients. PCR was positive for only one (0.5%) broncoalveolar lavage fluid of an adult patient and fifteen (7.5%) tracheal aspirates of pediatric patients. This study suggested that M. pneumoniae was more frequently detected in pediatric patients and PCR appears to have advantages over isolation, in terms of rapidity and sensitivity.
Central nervous system manifestations are probably the most frequent extrapulmonary complications of infections due to Mycoplasma pneumoniae, occur mostly in children. In this study, we attempted to isolate M. pneumoniae and to detect the organism by polymerase chain reaction (PCR) from cerebrospinal fluid samples (CSF) of pediatric patients. Of the 244 CSF samples, no M. pneumoniae was isolated. Six (2.5%) of the CSF samples were positive by PCR amplification. More effort are necessary to isolate the organism from CSF samples in order to ascertain the role of M. pneumoniae in causing neurological complications.