Highly specific and efficient primers for in-house multiplex PCR detection of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma hominis and Ureaplasma urealyticum.

BACKGROUND: Although sophisticated methodologies are available, the use of endpoint polymerase chain reaction (PCR) to detect 16S rDNA genes remains a good approach for estimating the incidence and prevalence of specific infections and for monitoring infections. Considering the importance of the early diagnosis of sexually transmitted infections (STIs), the development of a sensitive and affordable method for identifying pathogens in clinical samples is needed. Highly specific and efficient primers for a multiplex polymerase chain reaction (m-PCR) system were designed in silico to detect the 16S rDNA genes of four bacteria that cause genital infections, and the PCR method was developed. METHODS: The Genosensor Probe Designer (GPD) (version 1.0a) software was initially used to design highly specific and efficient primers for in-house m-PCR. Single-locus PCR reactions were performed and standardised, and then primers for each locus in turn were added individually in subsequent amplifications until m-PCR was achieved. Amplicons of the expected size were obtained from each of the four bacterial gene fragments. Finally, the analytical specificity and limits of detection were tested. RESULTS: Because they did not amplify any product from non-STI tested species, the primers were specific. The detection limits for the Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma hominis and Ureaplasma urealyticum primer sets were 5.12 × 10(5), 3.9 × 10(3), 61.19 × 10(6) and 6.37 × 10(5) copies of a DNA template, respectively. CONCLUSIONS: The methodology designed and standardised here could be applied satisfactorily for the simultaneous or individual detection of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma hominis and Ureaplasma urealyticum. This method is at least as efficient as other previously described methods; however, this method is more affordable for low-income countries.

[The importance of updating criteria of altered fasting glucose levels in Mexico]. / Importancia de la actualización en México del criterio de glucosa en ayuno alterada.

OBJECTIVE: To evaluate the impact of applying the 2003-ADA-proposed lower normality value of fasting glucose (FG) on impaired fasting glucose (IFG), prevalence and the agreement between diagnostic categories from ADA-2003 FG values and WHO two hours oral glucose tolerance test (OGTT) current criteria in a Mexican population with suspected diabetes. METHODS: A retrospective cross sectional study was undertaken. We analyzed fasting and 2 hours post load glucose values of 2062 patients and compared diagnostic categories on the basis of different criteria. RESULTS: Considering fasting values, prevalence of IFG changed from 17.7 % to 41.3 % when applying ADA-1997 or ADA-2003 criteria, respectively. Furthermore, based on their OGTT values (WHO-1999), 63 % the 852 IFG patients identified by ADA-2003 criteria were reclassified as having diabetes (26.1 %) or IGT (36.9 %). A heavy kappa test showed a moderate diagonal agreement of 0.43260 (CI 95 % = 0.43214-0.43305) between diagnostic categories from ADA-2003 with FG and OGTT values and WHO current criteria. CONCLUSIONS: The lower ADA-2003 criteria for IFG identifies a higher ratio of patients with IGT or DM.

Inclusion of educational messages in laboratory reports aids to complete the diagnostic workup of hyperglycemia.

OBJECTIVE: To evaluate whether educational messages regarding oral glucose tolerance test (OGTT) indications in laboratory reports increase the number of OGTTs appropriately requested. RESEARCH DESIGN AND METHODS: The following message was printed on the lab reports of individuals with a fasting plasma glucose (FPG) concentration between 5.5 and 6.9 mmol/l: "A FPG between 5.5 and 6.9 mmol/l is considered abnormal by the American Diabetes Association (impaired fasting glucose). An OGTT is recommended if the patient does not have a diagnosis of diabetes and suffers from conditions associated with an increased risk for having type 2 diabetes (i.e., overweight, high blood pressure, abnormal plasma lipids or family history of diabetes)." The number of educational messages printed was 81,099. RESULTS: The intervention resulted in a significant increase in the number of OGTTs requested, from 78 +/- 19 to 268 +/- 48 tests per month. It also resulted in a greater proportion of case subjects that had an abnormal OGTT result. CONCLUSIONS: Educational messages in laboratory reports aid in the diagnostic workup of hyperglycemia.