Physiological neutrophilia of pregnancy is not associated with a rise in plasma granulocyte colony-stimulating factor (G-CSF)

A patient with neutropenia and life-threatening infections secondary to T-γ lymphoproliferative disease, who did not respond to treatment with recombinant human G-CSF (filgrastim), was treated with filgrastim plus cyclosporine A (CyA). The patient achieved a good response in the absolute neutrophil count and subsequently required a dose reduction in the filgrastim. The patient was eventually discontinued from the CyA but continues on filgrastim alone. While on therapy, the large granular lymphocytes disappeared from the circulation and the beta-TCR rearrangement, which was present prior to beginning therapy, became undetectable. The patient had no significant toxicity to the CyA or the filgrastim and he has not experienced any serious infections or required hospitalization. Filgrastim has proven to be relatively nontoxic and of some benefit to patients with this disease and should probably be utilized first when treatment is necessary. However, if improvement is not observed, these findings suggest that a trial of the combination of CyA plus filgrastim may be beneficial.     

Using the polymerase chain reaction to maintain DNA probe inventories in clinical and diagnostic laboratories

Clinical and diagnostic DNA laboratories must maintain a large inventory of DNA probes for use in hybridization studies. The preparation of plasmid DNA and isolation of DNA fragments for use as probes in both expensive and time consuming. We present here a rapid and relatively inexpensive method of producing large amounts of DNA fragments from stocks, using the polymerase chain reaction (PCR). Our experience over the past year using this technique has been very positive and we believe many laboratories could benefit by employing such a labor-saving approach to maintaining DNA probes. The technique uses the bacteriophage M13 DNA sequencing primers to amplify cloned inserts contained in commonly used plasmid vectors. As examples, we illustrate the use of DNA produced in this manner as probes for linkage analysis of the fragile X syndrome and for detection of deletions in the Duchenne muscular dystrophy gene. We have also found that at least two probes can be amplified in the same PCR reaction, allowing the detection of two different restriction fragment length polymorphisms (RFLP) simultaneously. It should be possible for laboratories to devise strategies particular to their individual needs using more than one DNA probe produced in the same PCR reaction to detect RFLP's. Such strategies would need only to consider that the predicted alleles of the multiple polymorphisms do not migrate to the same position during electrophoresis. Stocks of single or multiple probes produced by the PCR could then be maintained for more rapid Southern analyses.     

Identification of Plasmid Containing Bacteria in an Activated Sludge Reactor

Samples taken from an activated sludge reactor used for the biodegradation of metal cutting fluids were studied for the presence of plasmid-containing bacteria. Twenty different bacterial isolates contained one or more plasmids. After numerous transfers of the isolates through LB broth, 55% of the plasmids were spontaneously lost as evidenced by agarose gel electrophoresis. Eighty percent of the isolates were resistant to one or more antibiotics, 65% resistant to two or more antibiotics, 40% resistant to three or more, 20% resistant to four or more and 5% resistant to five antibiotics. The isolates were also tested for their sensitivity to several common metals. This study has demonstrated that activated sludge is a natural reservoir for plasmid-containing bacteria involved in biodegradation of wastes.