Agreement between culture (L-J egg media) and multi gene PCR results using four set of primers.

<p><b>Abbreviations</b>: n: number of patients; %: percentage; H & E staining: Hematoxylin and Eosin; L-J egg medium: Lowenstein Jensen; AFB+Ve: Acid fast bacilli positive; Z-N staining: Ziehl-Neelsen's; PCR: Polymerase Chain Reaction; 19 kDa (131 bp):19 kDa antigen gene; TRC4 (173 bp): TRC4 element; MPB64 (240 bp): MPB64 antigen gene; 32 kDa (506 bp): 32 kDa protein (MTP-59 α-antigen) gene.</p

Laparoscopic/Hysteroscopic findings of infertile women suspected with female genital tuberculosis (FGTB) cases and control groups. Note

<p>: Some patients had more than one abnormal finding.</p

2.5% Agarose Gel Electrophoresis was carried out with Multi-gene/multi-primer PCR products.

<p>Lanes in the first row (118 to 136) were loaded with PCR products of FGTB patients; Lanes in the second row (C7 to C20) were loaded with PCR products of control patients; Lane +Ve Ctrl was loaded with positive Reference Strain (<i>M. tuberculosis</i>, ATCC 35836); Lane –Ve Ctrl was loaded with negative control (H₂O); Lane 50 bp marker was loaded with 50 base pair (bp) molecular weight ladder (50 bp size of product starts from the bottom side of gel and ends with 650 bp product on the top/upper side of the gel), The band corresponding to 131 bp was noted as 19 kDa antigen gene, 173 bp was noted as TRC4 element, 240 bp was noted as MPB64 gene, 506 bp was noted as 32 kDa protein/MPT59 α- antigen gene. Primer dimers were also noted at the bottom during the end of sample run.</p

Statistical significance tests between conventional and molecular methods (n = 302).

<p><b>Note</b>: “a” denotes binomial distribution used. Statistical analysis showed 0 cells (0.0%) have expected count less than 5 for all methods. Variation is observed for different methods in terms of the minimum expected counts (MEC), i.e. MEC for “b” is 34.77; “c” is 14.57; “d” is 28.48; “e” is 27.15; “f” is 42.05; “g” is 42.05; “h” is 47.02; and “i” is 41.06. Degree of freedom (df) is one (1) for all calculations. Data is considered statistically significant if p value is less than 0.05.</p><p><b>Abbreviations</b>: n: number of patients; %: percentage; H & E staining: Hematoxylin and Eosin; L-J egg medium: Lowenstein Jensen; AFB+Ve: Acid fast bacilli positive; Z-N staining: Ziehl-Neelsen's; PCR: Polymerase Chain Reaction; 19 kDa (131 bp):19 kDa antigen gene; TRC4 (173 bp): TRC4 element; MPB64 (240 bp): MPB64 antigen gene; 32 kDa (506 bp): 32 kDa protein (MTP-59 α-antigen) gene.</p

Demographic and clinical findings of FGTB cases and control groups (n = 302).

<p><b>Note</b>: Some patients had more than one abnormal finding; Data are presented as mean ± Standard Deviation (SD); n: number of patients; %: percentage.</p

Comparative analysis of methods for detection of FGTB cases-control groups (n = 302).

<p><b>Abbreviations</b>: Multi-gene PCR: Multi-gene/multi-primer Polymerase Chain Reaction; H & E staining: Hematoxylin and Eosin; L-J egg medium: Lowenstein-Jensen; AFB+Ve: Acid fast bacilli positive; Z-N staining: Ziehl-Neelsen's; PCR: Polymerase Chain Reaction; 19 kDa (131 bp): 19 kDa antigen gene; TRC4 (173 bp): TRC4 element; MPB64 (240 bp): MPB64 antigen gene; 32 kDa (506 bp): 32 kDa protein (MTP-59 α-antigen) gene. FGTB: female genital tuberculosis. The following terms were described based on demographic details, clinical symptoms, and hysteroscopic/laparoscopic findings of infertile women highly suspected with female genital tuberculosis (FGTB) and control women (without tuberculosis): <b>True Positive</b>: Infertile women with asymptomatic clinical presentations; radiologically may or may not be indicating tuberculosis (TB); positive diagnosis of TB on laparoscopic examinations; Indication of proliferative solid epitheliod granulomas, dense polymorphornuclear cells etc on histopathology; demonstration of acid fast bacilli in culture, Z-N staining of menstrual blood fluids, pelvic aspirated fluids and endo-ovarian tissue biopsies; detection of mycobacetrial DNA on multi gene PCR; and positive response to assisted reproductive technology (ART) after anti TB therapy among suspected FGTB cases. <b>True Negatives</b>: Fertile and healthy women; radiologically negative for TB; normal diagnosis (without TB) or absence of abnormal clinical findings on laparoscopic examinations; normal or negative finding on histopathology; absence of tubercle bacilli in culture and AFB negative on Z-N staining of menstrual blood or pelvic aspirated fluids or endo-ovarian tissue biopsies; absence of mycobacetrial DNA on multi gene PCR; Anti TB therapy is not required. <b>False Positive</b>: Detection of TB among control women (group without TB) by conventional and molecular methods; detection of TB among fertile and healthy women/negative findings of TB on laparoscopic examination. <b>False Negative</b>: Detection of negative TB by conventional and molecular methods among infertile asymptomatic women highly suspected of FGTB and among women diagnosed with positive TB on laparoscopic examinations.</p

Z-N staining of endo-ovarian tissue biopsy and cultures in the detection of acid fast bacilli (AFB).

<p>a) Red/pink colour rod like beaded structures were observed in the tissue biopsy; b) Red/pink colour rod like structures were observed on pale blue background in the cultures.</p

Histopathological examination of endo-ovarian tissue biopsy.

<p>a) Endometrial tissue biopsies from the surface resemble those of cervical microglandular hyperplasia with beaded spindle-cells; b) Beaded and malignant lymphoid cells were observed in ovarian tissue biopsies; c) Appearance of spindle-cells with superficial strips of positive endometrial glands and stroma; d) Lymphocytic infiltrations and initial stages of granulomatous were observed. <b>Note</b>: The microscopic studies were carried out with tissue biopsies and aspirated fluids containing tissue pieces. Thereafter, the slides were viewed under bright field (40X), Inverted microscope. The contrasts of the photographs are changed to improve the visibility.</p

Details of genes of <i>Mycobacterium tuberculosis</i> Complex.

<p>T: thymine; A: adenine; G: guanine; C: cytosine.</p

Diagnosis and statistical significance of FGTB infection among infertile women and HCW without TB.

<p><b>Note</b>: Statistical analysis showed 0 cells (0.0%) have an expected count less than 5 for all methods. Variation was observed for different methods in terms of the minimum expected counts, i.e. MEC for “a” is 33.45; “b” is 15.64; “c” is 30.55; “d” is 29.09 and “e” is 36.36. The “f” denotes the use of a binomial distribution. Degree of freedom (df) was one (1) for all calculations. Data were considered statistically significant if <i>p</i>-value was less than 0.05. Molecular analysis includes polymerase chain reaction (PCR) for 19kDa (131bp), 19kDa antigen gene; TRC4 (173bp), TRC4 element. AFB +Ve, AFB positive.</p><p>Diagnosis and statistical significance of FGTB infection among infertile women and HCW without TB.</p