Method for Reproducible Shipboard Segmented Flow Analysis Ammonium Measurement Using an In-House Reference Material for Quality Control

Ammonium is a fundamental nutrient for phytoplankton growth in seawater and is a key component of the microbial loop. Ammonium measured in parallel with other nutrients is crucial in understanding the small temporal scale changes in oceanographic ecology. Despite the importance of measuring ammonium at sea, owing to its lability, there is no consensus on the best method. The lack of availability of certified reference materials for ammonium in seawater also makes it difficult to assess the accuracy and reproducibility of ammonium measurements. In this study we present a modified segmented flow analysis method using ortho-phthaldialdehyde (OPA) with fluorescence detection to measure ammonium at sea together with four other macro-nutrients (nitrate, nitrite, silicate and phosphate) in near real time. An in-house ammonium quality control (QC) material was produced to improve the accuracy and repeatability of the measurement at sea. The QC was prepared following two different methods and stored in two types of containers. The suitability of the in-house QC’s as a reference material were assessed onboard the RV Investigator in 2018 during two oceanographic voyages, including one on the repeat SR03 CLIVAR transect. This paper describes the production and assessment of the in-house QC for ammonium in seawater, providing groundwork for creating a short-term stable ammonium reference material for sea going voyages. The uncertainty of this method of ammonium measurement was found to be 0.10 μmol/L at ammonium concentration of 1.0 μmol/L. Results show that preparation of the QC inside a laminar flow cabinet and directly into 10 mL polypropylene sample tubes just prior to the commencement of the voyage improved its stability.

Influence of dehydration on cryopreservation of Musa spp. germplasm

Cryopreservation is an important technique for the long-term storage of economically important plant germplasm. In this study, an efficient protocol was developed for the long-term conservation of seven economically important Musa taxa: M. acuminata Colla ssp. burmannica N.W. Simmonds, M. acuminata Colla ssp. zebrina (Van Houtte) R.E. Nasution, M. balbisiana Colla, M. basjoo Sieb., M. ornata W. Roxburgh (St. Lavender), M. velutina H. Wendl. et Drude (Velvet Pink Banana), and M. acuminata’ balbisiana. The seeds were dehydrated in a sterile laminar flow cabinet for different exposure times and then they were directly immersed in liquid nitrogen. The critical point was to support the initial germination of cryopreserved seeds and this was achieved by the excision of zygotic embryos after liquid nitrogen treatment that allowed the seed germination. The best moisture content for tolerance to cryopreservation ranged from 15.8% (M. acuminata ssp. zebrina) to 17.1% (M. ornata) and the maximum post-cryopreservation germination rates varied from 86.4% (M. velutina) to 55.0% (M. ornata). All seedlings derived from seeds germinated after cryopreservation were easily rooted and acclimated to greenhouse conditions.

POLYCLONAL OUTBREAK OF BLOODSTREAM INFECTIONS CAUSED BY Burkholderia cepacia COMPLEX IN HEMATOLOGY AND BONE MARROW TRANSPLANT OUTPATIENT UNITS

Aim: The objective was to describe an outbreak of bloodstream infections by Burkholderia cepacia complex (Bcc) in bone marrow transplant and hematology outpatients. Methods: On February 15, 2008 a Bcc outbreak was suspected. 24 cases were identified. Demographic and clinical data were evaluated. Environment and healthcare workers' (HCW) hands were cultured. Species were determined and typed. Reinforcement of hand hygiene, central venous catheter (CVC) care, infusion therapy, and maintenance of laminar flow cabinet were undertaken. 16 different HCWs had cared for the CVCs. Multi-dose heparin and saline were prepared on counter common to both units. Findings: 14 patients had B. multivorans (one patient had also B. cenopacia), six non-multivorans Bcc and one did not belong to Bcc. Clone A B. multivorans occurred in 12 patients (from Hematology); in 10 their CVC had been used on February 11/12. Environmental and HCW cultures were negative. All patients were treated with meropenem, and ceftazidime lock-therapy. Eight patients (30%) were hospitalized. No deaths occurred. After control measures (multidose vial for single patient; CVC lock with ceftazidime; cleaning of laminar flow cabinet; hand hygiene improvement; use of cabinet to store prepared medication), no new cases occurred. Conclusions: This polyclonal outbreak may be explained by a common source containing multiple species of Bcc, maybe the laminar flow cabinet common to both units. There may have been contamination by B. multivorans (clone A) of multi-dose vials.

Survival of human rhinovirus type 14 dried onto nonporous inanimate surfaces: effect of relative humidity and suspending medium

To study the survival of human rhinovirus 14 on environmental surfaces, each stainless steel disk (1 cm in diameter) was contaminated with 10 μL (about 105 plaque-forming units) of the virus suspended in either 1χ tryptose phosphate broth (TPB), 5 mg/mL of bovine mucin in normal saline, or undiluted human nasal discharge. The inoculum was dried in a laminar flow cabinet for 1 h under ambient conditions. The disks were then placed in a glass chamber (20 ± 1 °C) with the relative humidity at either low (20 ± 5%), medium (50 ± 5%), or high (80 ± 5%) level. At appropriate intervals, the disk to be tested was placed in 1 mL of tryptose phosphate broth and the eluate titrated in A-5 HeLa cells. When the virus was suspended in either tryptose phosphate broth, mucin, or the nasal discharge and subjected to initial drying, there was a 3.0 ± 1.0, 82.0 ± 6.7, and 89.0 ± 3.0% loss in virus infectivity, respectively. The half-life of the TPB-suspended virus was about 14 h at the high relative humidity as compared with < 2 h at the other two relative humidity levels. The half-lives for the mucin-suspended virus at the high, medium, and low relative humidity were 1.42, 0.55, and 0.24 h, respectively; the corresponding values for the nasal discharge suspended virus being 0.17, 0.25, and 0.09 h.

The use of laminar flow for obtaining germ-free mice

Hysterectomy in a laminar-flow cabinet affords greater economy of time and effort in obtaining germ-free mice than other methods. No contamination has occurred during the surgical procedure or in the subsequent transfer of the neonates into germ-free isolators.