gms | German Medical Science

Serological tests can be used to detect viral and bacterial antigens and antibodies (IgG and IgM) in a sample of blood, cerebrospinal fluid (CSF) or urine [1]. Serological testing is often used to help diagnose diseases, assess a patient’s immune and vaccination status, and control the spread of infection in a certain population [1], [2], [3]. Selecting the appropriate test can lead to a fast, accurate interpretation of the diagnostic findings, as well as proper and immediate treatment [1]. All tests have to meet standard operating procedures and should be validated or verified to comply with the laboratory’s quality management. Participation in external quality assurance (EQA) can ensure the accuracy of the diagnostic test results. EQA also allows a comparison of performance and results between different laboratories, the identification of problems associated with equipment, methods and materials, and the improvement of quality management. It also identifies training needs and monitors the role of training and its impact. EQA helps to assure physicians, patients and health authorities that the laboratory is producing reliable results. EQA participation is usually required for accreditation [2], [4]. This paper summarizes and discusses the results of bacteriological infection serology as part of proficiency testing trials conducted in 2017. The findings can help to improve the diagnosis of individual constellations and optimize the test systems used. This paper deals with a standardized form.

In 2017, 125,999 laboratories participated in two proficiency testing trials. 10,640 participants were from Germany and 1,450 were from other European countries. One trial was carried out in May and the other in November 2017 (Table 1 [Tab. 1]).

As the proficiency testing provider, the Society for Promoting Quality Assurance in Medical Laboratories e.V. (INSTAND e.V., Düsseldorf) in Germany sent unknown control samples (31, 32, 61 and 62) for each study group (310–334) to participating laboratories twice per year to test for one or more components present in the samples [5], [6]. All samples were prepared according to standard operating procedures [5], [6]. Samples 31 and 32 were sent to participating laboratories in May 2017, while samples 61 and 62 were sent in November 2017 [5], [6]. The control samples for Yersinia, Bordetella pertussis, Campylobacter, Mycoplasma pneumonia and Coxiella burnetii were only analyzed once that year [7], [8].

The results of all laboratories were compared to the assigned target value that was determined with the highest level of accuracy and precision using a reference measurement procedure in line with the guideline of the German Medical Association (RiLiBÄK). When a uniform target value could not be determined for the quantitative test results, the robust mean of all participants was established as the target value. With respect to the qualitative test results, either the mode of the results of the reference laboratories or the mode of the results of the participants was set as the target value [2], [7], [8].

Samples 31, 32, 61 and 62 originated from clinically healthy blood donors.

For the tests conducted in May, the mode of the results of all participants was set as the target value for the qualitative test results, while the mode of the results of the reference laboratories was established as the target value for the semi-quantitative test results. For the November tests, the mode of the results of the reference laboratories was set as the target value for the qualitative test results, while the robust mean of all participants was established as the target value for the semi-quantitative test results [7], [8]. The results are listed in Table 2 [Tab. 2].

Antibody detection has no diagnostic relevance in determining a tetanus infection, but it can be used to assess an individual’s immune or vaccination status for tetanus toxoid [1]. Results of this trial show that samples 31, 61 and 62 had tetanus toxoid antibody titers ranging from 1.4 to 2.9 IU/ml, suggesting protective immunity against tetanus toxoid; however, a booster vaccination would be needed in 5 to 10 years to achieve long-term protection. The level of antibodies in sample 32 was 0.9 IU/ml, suggesting that there was protective immunity; however, a booster vaccination would be needed in 2 to 5 years to provide long-term protection. It is important to note that vaccinations should primarily be done in accordance with the recommendations of the German Standing Committee on Vaccination (STIKO) and not based simply on measured antibody levels [3]. Pass rates for ELISA tests were 100% for the qualitative results and 81.5–96.8% for the quantitative results. The pass rate for the overall diagnostic results was between 98.5% and 99.3%.

Samples 32 and 62 originated from healthy blood donors without clinical evidence of syphilis. Sample 31 was taken from a patient with a known syphilis infection that had been sufficiently treated in the past. Sample 61 was donated during blood donor screening by an individual that had been treated for a syphilis infection several years ago.

For the qualitative test results, the mode of the results of the reference laboratories was set as the target value. In the case of the semi-quantitative test results, the mode of the results of all participants was set as the target value for the May tests, while the results of the reference laboratories were stipulated as the target value for November [7], [8]. The results are listed in Table 3 [Tab. 3].

Samples 32 and 62 showed no clinical or serological evidence of a syphilis infection. In the positive sample 31 (target value (modal): TPPA: 320, VDRL: negative, IgM-FTA-ABS: negative), the results clearly indicated a past infection without the need for further treatment because the IgM-FTA-ABS test was negative. The overall pass rate of all test methods in sample 31 was 77.2%. The positive sample 61 (target value: TPPA: 640, polyval. ELISA: positive, IgG-ELISA: positive, VDRL: negative, FTA-ABS-IgG: 80, FTA-ABS-IgM and IgM-ELISA: negative) indicated a syphilis infection and achieved an overall pass rate for all test methods of 86.9%. The distribution of the immunoblot bands for the positive samples 31 (Figure 1 [Fig. 1], Figure 2 [Fig. 2]) and 61 (Figure 3 [Fig. 3]) are shown below [7], [8]. The pass rates for the overall diagnostic results of the negative samples 32 and 62 were between 96.4% and 98.8%, whereas the pass rates of the positive samples 31 and 61 were between 77.2% and 86.9%.

Samples 31, 32, 61 and 62 originated from clinically healthy blood donors.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results, while for the semi-quantitative results, the mode of the results of all participants was set as the target value [7], [8]. The results are listed in Table 4 [Tab. 4].

Samples 32 and 61 showed no serological evidence of a C. trachomatis infection. In sample 31, IgG antibodies by ELISA C. spp and blot as well as IgA-ELISA C. spp were detected, indicating an infection with C. trachomatis. In sample 62, IgG and IgA antibodies were detected, suggesting an infection with C. trachomatis. In sample 61, the lowest pass rate was 50% for the qualitative detection of IgG antibodies by immunoblot. In sample 31, the lowest pass rates for the qualitative detection of IgG antibodies by immunoblot and ELISA were 50% and 50.9% respectively. The pass rates for the overall diagnostic results of the negative samples (samples 32 and 61) were between 76.1% and 98.8%, while the pass rates for the positive samples (samples 31 and 62) were between 86.7% and 98.4%.

Samples 31, 32, 61 and 62 originated from clinically healthy blood donors.

For the qualitative test results, the mode of the results of all participants was set as the target value. In the case of the semi-quantitative test results, the mode of the results of the reference laboratories was established as the target value for the tests conducted in May, while the mode of the results of all participants was stipulated as the target value for November [7], [8]. The results are presented in Table 5 [Tab. 5].

Samples 31 and 62 showed no clinical or serological evidence of C. trachomatis, while samples 32 and 61 tested positive for the pathogen, indicating a C. trachomatis infection. The pass rate of all samples for the overall clinical diagnostic results was 100%.

Samples 31, 32, 61 and 62 were taken from clinically healthy blood donors.

For the tests conducted in May, the results of the reference laboratories was set as the target value for the qualitative results, while for the semi-quantitative test results, the mode of the results of the reference laboratories was set as the target value. For November, the mode of the results of the reference laboratories was set as the target value for the qualitative test results, while for the semi-quantitative results, the mode of the results of all participants was set as the target value [7], [8]. The results and pass rates are shown in Table 6 [Tab. 6].

In samples 32, 61 and 62, no evidence of an infection with C. pneumoniae was detected, while the test results for the tested pathogen in sample 31 were positive. In sample 31, IgG was detected, with borderline positive results for IgA antibodies, indicating an infection with C. pneumoniae. All samples had qualitative results of 84.6–100% for all ELISA tests. All MIFT tests were between 66.7 and 100% with respect to the qualitative results and in a range of 64.7–100% for the quantitative results. The pass rates for overall diagnostic results of the negative samples 32, 61 and 62 were between 96.8% and 100%, while for the positive sample 31, the pass rate was 98.2%.

Sample 31 was obtained from a healthy blood donor. Sample 32 was donated by a patient with gastroenteritis and arthralgia in his recent medical history.

For the tests conducted in May, the mode of the results of all participants was set as the target value for the qualitative test results, while for the semi-quantitative test results, the mode of the results of the reference laboratories was set as the target value. However, in November, the mode of the results of the reference laboratories was set as the target value for the qualitative test results, while the results of the reference laboratories were stipulated as the target value of the semi-quantitative results [7], [8]. The results are listed in Table 7 [Tab. 7].

Samples 31, 32, 61 and 62 were taken from clinically healthy blood donors.

The mode of the results of all participants was established as the target value for the qualitative test results [7], [8]. The results are listed in Table 5 [Tab. 5].

Samples 32 and 62 showed no serological evidence of C. trachomatis, while samples 31 and 61 tested positive for the pathogen, indicating an infection with C. trachomatis. The pass rates for the overall diagnostic results of the negative samples 32 and 62 were between 95.7% and 100% and thus in line with previous years, while the pass rate for the positive samples 31 and 61 was 100%.

Samples 61 and 62 were donated by healthy blood donors without evidence of any respiratory infections in their recent medical history.

For the qualitative test results, the mode of the results of the reference laboratories was set as the target value, while for the semi-quantitative test results, the robust mean of all participants was established as the target value [7], [8]. The results are listed in Table 8 [Tab. 8].

Samples 61 and 62 were donated by healthy pre-immunized blood donors, while samples 31 and 32 originated from clinically healthy blood donors.

In the case of the semi-quantitative test results, the robust mean of all participants was established as the target value. For the qualitative test results, the mode of the results of the reference laboratories was set as the target value for the May tests, while for November, the mode of the results of all participants was set as the target value [7], [8]. The results are shown in Table 9 [Tab. 9].

Antibody detection against the diphtheria toxin and toxoid (DT) is not suitable for identifying an acute case of diphtheria; it can only be used to assess the immune and vaccination status for diphtheria [3]. Findings suggested that protective immunity did exist in samples 61 and 62, while in sample 31, there was no protective immunity, therefore a booster vaccination was recommended. The titer level (0.922 IU/ml) of sample 32 indicated protective active immunity, however, a booster vaccination would be needed to provide long-term protection. Vaccination recommendations should primarily be made following STIKO recommendations [3]. ELISA tests had qualitative results of 100% and quantitative results of 88.1–97.6%. The pass rates for the overall diagnostic results were in the range of 96.0–100%.

Samples 31 and 32 originated from clinically healthy blood donors.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results. In the case of the semi-quantitative test results, the mode of the results of all participants was set as the target value for the May tests; however, the robust mean of all participants was established as the target value [7], [8] for November. The results are listed in Table 10 [Tab. 10].

Specific IgG, IgM and IgA antibodies were detected using the commercially available ELISA, immunoblot and IFT tests. No evidence of an infection with C. jejuni was detected in sample 32, while sample 31 tested positive for IgG antibodies against the tested pathogen, indicating a Campylobacter infection. In sample 31, qualitative and quantitative results for ELISA were 100% for IgG, IgM and IgA antibodies, while in sample 32 they ranged from 91.7–100%. The pass rate for the overall diagnostic results of sample 32 was 96.52%, while it was 86.8% for sample 31.

Samples 31, 32, 61 and 62 were donated by healthy blood donors.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results, while the robust mean of all participants was established as the target value for the semi-quantitative test results [7], [8]. The results are presented in Table 11 [Tab. 11].

The findings of samples 32 and 66 indicated the likelihood of systemic inflammatory reaction (sepsis), while the findings of samples 31 and 61 indicated that systemic infection is unlikely. The qualitative result in all four samples was 100%. The quantitative test results in samples 31 and 61 were 100%, while they were 92.8% in sample 32, and 95.6% in sample 62. The semi-quantitative results in samples 31 and 32 were 100%, while they were 93.8% in samples 61 and 62. The overall diagnostic evaluation of all four samples was between 89.5 and 100%.

Samples 31 and 32 were donated by healthy blood donors.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results, while in the case of the semi-quantitative test results, the robust mean of all participants was established as the target value [7], [8]. The results are displayed in Table 12 [Tab. 12].

In the qualitative results, titers of streptococcal antibodies above the cut-off value (200 IU/ml) indicated an infection with Streptococcus. Titers between 200 and 400 indicated a past or recent infection [5]. A much higher titer occurs when there is severe infection or an acute secondary disease. The latex agglutination method used to detect Streptococcus-O-lysine antibodies had a qualitative and quantitative test result of 33.3% in sample 31, while in sample 32, the quantitative test result was 66.7%. The overall pass rate of Streptococcus-O-lysine antibody detection was 100%, while the overall pass rate of streptodornase detection was between 33.3% and 100%.

Samples 31 and 32 were taken from clinically healthy blood donors.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results, while in the case of the semi-quantitative test results, the robust mean of all participants was established as the target value [7], [8]. The results are indicated in Table 13 [Tab. 13].

The qualitative test results for both samples for all methods were 100%. The qualitative and quantitative test results for sample 31 for method 1 was 50%, while the quantitative test result for sample 32 for method 1 was 50%.

Samples 61 and 62 originated from patients with several known respiratory infections in their recent medical history.

The mode of the results for all participants was set as the target value for the qualitative and semi-quantitative test results [7], [8]. The results are presented in Table 14 [Tab. 14].

Evidence of an infection with M. pneumonia was detected in both samples. In both samples both the reference laboratories and most participants found variable results and weak IgG and IgA seroreactivity [7], [8]. The pass rate for the overall diagnostic results was 100% for sample 61 and 82.7% for sample 62.

Sample 61 was donated by a healthy blood donor who showed no evidence of a recent infection. Sample 62 was donated by a patient a few months after having acute C. burnetii pneumonia.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results, while the mode of the results of all participants was set as the target value for the semi-quantitative test results [7], [8]. The results are shown in Table 15 [Tab. 15].

Sample 61 tested negative for C. burnetii, while sample 62 tested positive. Sample 61 exhibited IgG phase I IFT titers of 5120 (median), IgG phase II IFT titers of 2560 (median) as well as weakly reactive IgM and IgA results. These results suggested a relatively recent case of pneumonia. Most participants and expert laboratories made variable clinical comments as to whether the test constellation should be interpreted as an acute or chronic Coxiella infection [7], [8].

The pass rate for the overall diagnostic results of sample 61 was 100%, while the pass rate of sample 62 was 89.0%.

Samples 31, 32, 61 and 62 originated from clinically healthy blood donors.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results, while the results of the reference laboratories were stipulated as the target value of the semi-quantitative results [7], [8]. The results are listed in Table 16 [Tab. 16].

No evidence of a Salmonella infection was detected in all samples. The pass rates for all samples for the overall diagnostic results were between 94.4% and 98.4%.

Samples 31 and 61 originated from a healthy blood donor without evidence of a tick bite or clinical Lyme borreliosis in his medical history. Sample 32 was donated by a patient with past Lyme arthritis treated several years ago and confirmed by culture and PCR. Sample 62 was donated two years after the infection by a patient with successfully treated Lyme arthritis.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results [7], [8]. The results are listed in Table 17 [Tab. 17].

No evidence of an infection with B. burgdorferi was detected in samples 31 and 61, while pathogens were detected in samples 32 and 62, indicating an infection with B. burgdorferi. Samples 32 and 62 showed high IgG antibody titers together with borderline reactive IgM test results, suggesting a late phase of the borrelia-specific immune response. Figure 4 [Fig. 4] and Figure 5 [Fig. 5] show the distribution of specific IgG and IgM borrelia immunoblot bands for sample 32 [7], [8]. The distribution of the immunoblot bands reported for the positive sample 62 is depicted in Figure 6 [Fig. 6].

The pass rates for the overall diagnostic results of the negative samples 31 and 61 were between 98.2% and 98.7%, while for the positive samples 32 and 62 they were between 82.5% and 94.8%.

Samples 31, 32 and 61 originated from clinically healthy blood donors. Sample 62 was taken from a helicobacter-positive patient shortly after finishing eradication therapy.

The mode of the results of the reference laboratories was set as the target value for the qualitative test results. The results are shown in Table 18 [Tab. 18].

There was no evidence of an infection with H. pylori in samples 32 and 61. Samples 31 and 62 showed positive IgG and IgA antibody reactivity by ELISA and immunoblot, indicating an infection or colonization with H. pylori. In sample 61, the qualitative result for IgA blot was 60%, while for IgG blot it was 71.4%. The pass rates for the overall diagnostic results of the positive samples 31 and 62 were between 91.3% and 93.0%, while for the negative samples 61 and 32 they were between 82.5% and 95.7%.

EQA can be used as a tool in improving diagnostic processes in laboratories. It leads to more reliable and prompt diagnostic results, which sustain the quality and efficiency of patient care. The results of the EQA presented in this report generally show moderate to very good diagnostic quality. However, some special comments on serology need to be made:

Regarding the Treponema pallidum serology (311), sample 31 had low pass rates of 66.7% for the qualitative detection of IgM antibodies by immunoblot and 73.9% for the detection of IgG antibodies by FTA-ABS. These contributed to the poor overall pass rate of 77.2% for all test methods. Generally, laboratory results in 2017 showed better pass rates compared to the results in 2016. In the case of Chlamydia trachomatis serology (312), the overall pass rate was between 50% and 100% for all tests in sample 31. A pass rate of 50% was observed for the qualitative detection of IgG antibodies by immunoblot. Next, pass rates of 50.9% and 63.6% were observed for the qualitative detection of IgG and IgM antibodies respectively, both detected by ELISA in the above-mentioned sample. In sample 61, the lowest pass rate of 50% was observed for the qualitative detection of IgG antibodies by immunoblot, while in sample 62 the lowest pass rate was 75% for the same method. With regard to Chlamydia pneumonia serology (314), the overall pass rate was between 64.7% and 100% for all tests in sample 31. A pass rate of 66.7% was observed for the qualitative detection and 64.7% for the quantitative detection of IgM antibodies by MIFT. A pass rate of 77.8% was observed for the qualitative detection of IgA antibodies, also by MIFT, and a pass rate of 66.7% was recorded for the qualitative detection of IgG antibodies by immunoblot in the above-mentioned sample. In sample 32, the lowest pass rate of 77.8% was observed for the qualitative detection of IgM antibodies by MIFT. Generally, the pass rates were lower in 2017 compared to the results in 2016. In the case of Yersinia antibody serology (315), the lowest pass rate of 76.9% was observed for the qualitative detection of IgG antibodies by immunoblot in sample 31. Sample 32 recorded the lowest pass rate of 70% for the qualitative detection of IgM antibodies by immunoblot. In the case of Campylobacter serology (319), the lowest pass rate of 64.7% was observed for qualitative and 72.2% for quantitative detection by CFT in sample 31. In the same sample, a pass rate of 75% was detected for the qualitative detection of IgA by immunoblot. Sample 32 recorded the lowest pass rate of 75% for the qualitative detection of IgG antibodies by immunoblot. With respect to Streptococci serology (321), the pass rate was 33.3% for qualitative and quantitative detection by latex agglutination in sample 31, while in sample 32, a pass rate of 66.7% for quantitative detection by latex agglutination was observed. Generally, the pass rates were lower in 2017 than in 2016. Regarding rheumatoid factor serology (323), the pass rate was 50% for qualitative and quantitative detection using the nephelometry method in sample 31, while a pass rate of 50% for quantitative detection by the same method mentioned above was observed in sample 32. With regard to Mycoplasma pneumonia serology (324), the pass rate was 63.6% and 66.7% for qualitative and quantitative detection respectively, using PHA in sample 62. In the same sample, the pass rate was 68.9% for the quantitative detection of IgA antibodies by ELISA. With respect to the Coxiella burnetii serology (325), the overall pass rate was 100% in the diagnostic assessment of sample 61. The lowest pass rate for quantitative detection of IgA by IFT was 63.6%, showing poorer laboratory results than the pass rate in 2016. In the case of the Borrelia burgdorferi serology (332), both samples 31 and 32 showed the lowest pass rate of 75% in the qualitative detection by line immunoblot. In the diagnostic assessment of sample 31, an overall pass rate of 98.2% was recorded, while in sample 32, it was 82.5%. In the Helicobacter pylori serology (334), the overall pass rate for sample 61 was between 60% and 100%. In terms of the qualitative result, the pass rate was 71.4% for IgG detection and 60% for IgA detection, both by immunoblot.

The above-mentioned findings and comments show, as in previous years, the need for further improvement in certain diagnostic procedures. This will achieve more standardized and higher quality testing in Germany to ensure even better diagnostic test results in the routine clinical setting. This will maintain quality and efficiency in patient care.

The authors declare that they have no competing interests.