Megan L. Lloyd

Microbiology Australia, bringing Microbiologists together

Microbiology is increasingly being incorporated into secondary school biology curricula, an important component of science technology engineering and mathematics (STEM) training. The microbiological competency of teachers and the availability of skilled laboratory technicians that understand the risks involved and the reasoning behind restrictions to protocols is vital for the safe execution of microbiology experiments in secondary schools. The addition of gene-modification experiments to microbiology curricula increases the complexity of experimental set-up and potentially exacerbates existing deficiencies in laboratory technique and disinfection practice. There is currently no targeted national program to up-skill school science teachers or laboratory technicians, which can leave these staff in vulnerable positions if they are asked to co-ordinate experiments that are beyond their expertise. It is important that such training is available, preferably as part of well-planned professional development and that continued online support is provided to answer questions swiftly and assist STEM school staff to conduct microbiology experiments safely. Improved integration between universities and high schools can be beneficial in this regard, not only in the safe teaching of microbiology in schools, but also in collaborative links formed between secondary schools and universities.

Since murine cytomegalovirus (MCMV) was first described in 1954, it has been used to model human cytomegalovirus (HCMV) diseases. MCMV is a natural pathogen of mice that is present in wild mice populations and has been associated with diseases such as myocarditis. The species-specific nature of HCMV restricts most research to cell culture-based studies or to the investigation of non-invasive clinical samples, which may not be ideal for the study of disseminated disease. Initial MCMV research used a salivary gland-propagated virus administered via different routes of inoculation into a variety of mouse strains. This revealed that the genetic background of the laboratory mice affected the severity of disease and altered the extent of subsequent pathology. The advent of genetically modified mice and viruses has allowed new aspects of disease to be modeled and the opportunistic nature of HCMV infection to be confirmed. This review describes the different ways that MCMV has been used to model HCMV diseases and explores the continuing difficulty faced by researchers attempting to model HCMV congenital cytomegalovirus disease using the mouse model.

Microbiology Australia, bringing Microbiologists together

Human cytomegalovirus (CMV) is often transmitted through saliva. The salivary gland is a site of CMV replication and saliva can be used to diagnose congenital CMV infections. CMV replication is monitored in whole blood or plasma in renal transplant recipients (RTR) and associates with clinical disease. However, these assays may not detect replication in the salivary gland and there is little data linking detection in saliva with systemic infection and clinical sequelae. RTR (n = 82) were recruited > 2 years after transplantation. An in-house quantitative PCR assay was used to detect CMV UL54 in saliva samples. CMV DNA was sought in plasma using a commercial assay. Vascular health was predicted using flow mediated dilatation (FMD) and plasma biomarkers. CMV-reactive antibodies were quantified by ELISA and circulating CMV-specific T-cells by an interferon-γ ELISpot assay. Vδ2⁻ γδ T-cells were detected using multicolor flow cytometry reflecting population expansion after CMV infection. The presence of CMV DNA in saliva and plasma associated with plasma levels of antibodies reactive with CMV gB and with populations of circulating Vδ2⁻ γδ T -cells (p < 0.01). T-cells reactive to CMV immediate early (IE)-1 protein were generally lower in patients with CMV DNA in saliva or plasma, but the level of significance varied (p = 0.02–0.16). Additionally, CMV DNA in saliva or plasma associated weakly with impaired FMD (p = 0.06–0.09). The data suggest that CMV detected in saliva reflects systemic infections in adult RTR.

Introduction:S. aureus is one of the most prevalent potential pathogenic bacteria found in DHM. Some strains produce heat stable enterotoxins that are able to survive pasteurization. These enterotoxins have been associated with gastritis and potentially necrotizing enterocolitis in preterm infants. The aim of this study is to assess the impact of different storage temperatures on Staphylococcus aureus growth and enterotoxin A and B production in raw and Holder pasteurized donor human milk (DHM). Materials and Methods: Raw and pasteurized DHM were inoculated with enterotoxin A and B producing S. aureus. Samples were incubated at 4°C (10 days), 21°C, and 37°C (18 hours). Bacterial growth and enterotoxin A and B were quantified. Results:S. aureus count increased in pasteurized DHM. Bacterial count decreased in the raw milk when incubated at 21°C and 4°C and slightly increased when incubated at 37°C. Enterotoxins A and B were only detected in pasteurized DHM at 37°C from 9 hours onward. Conclusion: This study showed that raw milk is capable of suppressing S. aureus growth compared to pasteurized DHM. It also provides a measure of assurance of the safety of raw and pasteurized DHM when refrigerated or left at room temperature for a few hours.

Background: Some strains of Staphylococcus aureus can produce heat-stable enterotoxins that have been associated with gastritis and potentially necrotizing enterocolitis in preterm infants. Objectives/Hypothesis: To assess the impact of different storage temperatures on S. aureus growth and enterotoxin production in raw, Holder-pasteurized (HP) and ultraviolet-C (UV-C)-treated donated human milk (DHM). Materials and Methods: The milk samples from individual donors were pooled and divided into four equal portions. One portion was HP, the second was UV-C treated, the third was not treated, and the fourth was UV-C treated after being spiked with S. aureus. All samples were incubated at 37°C (18 hours) and 4°C (14 days). Bacterial colony count, enterotoxin A and B, and immune proteins were quantified. Results: At 37°C, the colony count increased in HP DHM and decreased in raw and UV-C-treated DHM. At 4°C, colony counts in HP DHM reduced and were not detected in raw and UV-C-treated DHM from day 8 of incubation. No bacteria were detected in samples that were inoculated before UV-C treatment. Enterotoxin A was only detected in HP-DHM at 37°C from the 9th hour onward. Enterotoxin B was detected in one sample at the 15th hour. Immune protein concentrations were similar in raw and UV-C DHM, and were reduced in the HP DHM. Conclusion: UV-C-treated milk reduces S. aureus growth with similar kinetics to raw milk making it a promising emerging technique to eliminate bacteria while retaining essential immune proteins in DHM.

Preterm infants whose mothers are unable to produce sufficient breast milk are increasingly being supplemented with pasteurised donor human milk (PDHM) instead of commercial preterm infant formula. Concerns have been raised that this practice can result in reduced growth. This retrospective clinical audit collected data from the medical records of a cohort of preterm infants (≤30 weeks gestational age) receiving either ≥28 d of PDHM (n 53) or ≥28 d of their mother’s own milk (MOM, n 43) with standard fortification supplied to both groups during admission. Weight growth velocity was assessed from regained birth weight to 34+1 weeks’ postmenstrual age (PMA); and weight, length and head circumference were compared at discharge and 12 months (corrected age). At 34+1 weeks’ PMA, the weight growth velocity (g/kg per d) was significantly lower in the PDHM group (15·4 g/kg per d, 95 % CI 14·6, 16·1) compared with the MOM group (16·9 g/kg per d, 95 % CI 16·1, 17·7, P=0·007). However, the increase was still within clinically acceptable limits (>15 g/kg per d) and no significant difference was observed in the weight between the two groups. There was no significant difference in weight between the groups at discharge or at the 12-month corrected gestational age review. Although we demonstrated a significant reduction in the weight growth velocity of preterm infants receiving PDHM at 34 weeks’ PMA, this difference is not present at discharge, suggesting that the growth deficit is reduced by supplementation before discharge.

The American Society for Microbiology's curricular guidelines for Introductory Microbiology highlighted key laboratory skills in the isolation, visualization, and identification of microorganisms as core learning objectives in the discipline. Since the publication of these guidelines in 2012, there has been a paucity of diagnostic assessment tools in the literature that can be used to assess competencies in the microbiology laboratory. This project aimed to establish a laboratory competency examination for introductory microbiology, with tasks specifically aligned to laboratory skills and learning outcomes outlined in curricular guidelines for microbiology. A Laboratory Competency Examination assessing student skills in light microscopy, Gram-staining, pure culture, aseptic technique, serial dilution, dilution calculations, and pipetting was developed at The University of Queensland, Australia. The Laboratory Competency Examination was field-tested in a large introductory microbiology subject (∼400 students), and student performance and learning gains data was collected from 2016–2017 to evaluate the validity of the assessment. The resulting laboratory assessment is presented as an endpoint diagnostic tool for assessing laboratory competency that can be readily adapted towards different educational contexts.