Mismatch amplification mutation assay-polymerase chain reaction: A method of detecting fluoroquinolone resistance mechanism in bacterial pathogens

The mismatch amplification assay is a modified version of polymerase chain reaction (PCR) that permits specific amplification of gene sequences with single base pair change. The basis of the technique relies on primer designing. The single nucleotide mismatch at the 3' proximity of the reverse oligonucleotide primer makes Taq DNA polymerase unable to carry out extension process. Thus, the primers produce a PCR fragment in the wild type, whereas it is not possible to yield a product with a mutation at the site covered by the mismatch positions on the mismatch amplification mutation assay (MAMA) primer from any gene. The technique offers several advantages over other molecular methods, such as PCR-restriction fragment length polymorphism (RFLP) and oligonucleotide hybridization, which is routinely used in the detection of known point mutations. Since multiple point mutations in the quinolone resistance determining region play a major role in high-level fluoroquinolone resistance in Gram-negative bacteria, the MAMA-PCR technique is preferred for detecting these mutations over PCR-RFLP and sequencing technology.

Serology and immunoglobulin profile in rheumatoid arthritis.

One hundred and twenty cases of clinically diagnosed rheumatoid arthritis, 80 non-rheumatoid cases suffering from various other diseases and 40 healthy individuals were investigated for the presence of rheumatoid factor, quantitation of serum immunoglobulin, demonstration of ANA and LE cell phenomenon. Microlatex agglutination test of serum for rheumatoid factor showed 56.6% positivity in rheumatoid group and 3.7% positivity in non-rheumatoid group. All three serum immunoglobulins (IgG, IgM, IgA) were raised in serum in significant titre in cases of rheumatoid arthritis, whereas only IgA lever was elevated in the group of non-rheumatoid diseases. ANA and LE cell phenomenon were observed in 11.7% and 4.4% cases of rheumatoid arthritis who had severe underlying disease. In non-rheumatoid group, only one of 6 cases of systemic lupus erythematosus showed rheumatoid factor and that too in an insignificant titre (less than 1:20). Synovium and synovial fluid contained plenty of plasma cells and lymphocytes. It has been observed that RF appears first in synovial fluid and it may take several months to a year to attain detectable level in serum.

Toxoplasmosis in women of child bearing age and infant follow up after in-utero treatment.

A total of 540 women (including 70 pregnant cases) of child bearing age with bad obstetrical history were tested serologically for anti-toxoplasma antibody using microlatex agglutination test. Forty two women including 5 cases of pregnancy were found to be seropositive in a titre of 1:32 or more. Maximum prevalence (10.2%) and highest titer of anti-toxoplasma antibodies were observed in women of 35-42 years age group. The overall prevalence of toxoplasmosis in these women was 7.7%, whereas it was 7.1% in pregnant women. Further studies are needed to estimate the exact rate of prevalence of infection. Of the 70 pregnant women, 5 were seropositive and two of them acquired infection during pregnancy which was detected by IgM immunosorbent assay. Seropositive pregnant women were treated using combined regimen of sulfadiazine and pyrimethamine. Four infected women with pregnancy were followed up and one did not turn up subsequently. There was spontaneous abortion in one case and in 3 other cases full term normal babies were delivered. Incidence of toxoplasmosis in women is low because of infrequent and uncommon practices of ingesting undercooked or uncooked food stuff specially meat by a substantial number of the population surveyed.

Neonatal candidemia.

Fifty five clinically diagnosed cases of neonatal septicaemia of the nursery ward, Medical College Hospital Calcutta were subjected to blood culture on two consecutive days in all cases. Bacterial isolates were found in 34 (61.8%) cases as pure growth and candida species from 9 (16.4%) as pure culture. Remaining 12 samples did not yield any growth. Species distribution of candida showed C. albicans 6, C. parapsilosis 2, and C. guillermondii 1. Asphyxia neonatorum was the common feature in all cases of candidaemia. All neonates with candidal infection were low-birth-weight and premature. C. albicans, the predominant species (66.6%) recovered, was responsible for a localised outbreak of infection in the nursery.