Use of Shell-Vial Cell Culture Assay for Isolation of Bacteria from Clinical Specimens: 13 Years of Experience

Abstract

The shell-vial culture assay is performed routinely in our laboratory. Recently we revisited our experience of using the shell-vial culture assay for the isolation of microorganisms from various clinical samples. Over a 13-year period, we have isolated 580 bacterial strains (5%) from 11,083 clinical samples tested. Over the same period, 285 isolates of rickettsiae, bartonellae, or Coxiella burnetii were cultured from a total of 7,102 samples tested. These isolates include 55 Rickettsia sp. isolates, 95 Coxiella burnetii isolates, and 135 Bartonella sp. isolates. Based on our experience with the growth of fastidious microorganisms, we have used a centrifugation shell-vial technique called JNSP, for “je ne sais pas” (“I don't know [what I am growing]”) for the isolation of other microorganisms. A total of 173 isolates were cultured from the 3,861 clinical samples tested using the JNSP method. Of these, 40 isolates had not been grown before on usual axenic medium. These include 2 Staphylococcus aureus isolates, 7 isolates of Streptococcus sp. and related genera, 6 Mycobacterium sp. isolates, 1 Nocardia asteroides isolate, 1 Actinomyces sp. isolate, 1 Brucella melitensis isolate, 2 Francisella tularensis isolates, 1 Mycoplasma pneumoniae isolate, and 1 Legionella pneumophila isolate. Using this protocol, we have also cultured intracellular bacteria such as Chlamydia trachomatis and we have performed the first culture and establishment of Trophyrema whipplei . Applied in our laboratory, the shell-vial culture generally exhibits a low rate of success. However, in some cases, this technique allowed microbial diagnosis when classical agar procedure and PCR were negative.