Internal validation of STRmix™ – A multi laboratory response to PCAST

We report a large compilation of the internal validations of the probabilistic genotyping software STRmix™. Thirty one laboratories contributed data resulting in 2825 mixtures comprising three to six donors and a wide range of multiplex, equipment, mixture proportions and templates. Previously reported trends in the LR were confirmed including less discriminatory LRs occurring both for donors and non-donors at low template (for the donor in question) and at high contributor number. We were unable to isolate an effect of allelic sharing. Any apparent effect appears to be largely confounded with increased contributor number

Particular genomic and virulence traits associated with preterm infant-derived toxigenic Clostridium perfringens strains

Clostridium perfringens is an anaerobic toxin-producing bacterium associated with intestinal diseases, particularly in neonatal humans and animals. Infant gut microbiome studies have recently indicated a link between C. perfringens and the preterm infant disease necrotizing enterocolitis (NEC), with specific NEC cases associated with overabundant C. perfringens termed C. perfringens-associated NEC (CPA-NEC). In the present study, we carried out whole-genome sequencing of 272 C. perfringens isolates from 70 infants across 5 hospitals in the United Kingdom. In this retrospective analysis, we performed in-depth genomic analyses (virulence profiling, strain tracking and plasmid analysis) and experimentally characterized pathogenic traits of 31 strains, including 4 from CPA-NEC patients. We found that the gene encoding toxin perfringolysin O, pfoA, was largely deficient in a human-derived hypovirulent lineage, as well as certain colonization factors, in contrast to typical pfoA-encoding virulent lineages. We determined that infant-associated pfoA + strains caused significantly more cellular damage than pfoA − strains in vitro, and further confirmed this virulence trait in vivo using an oral-challenge C57BL/6 murine model. These findings suggest both the importance of pfoA + C. perfringens as a gut pathogen in preterm infants and areas for further investigation, including potential intervention and therapeutic strategies

The prevalence of mixed DNA profiles in fingernail samples taken from couples who co-habit using autosomal and Y-STRs

Physical contact can result in the transfer of DNA from one individual to another. In cases of sexual offence this can often be by means of an aggressive or Sexual act. Biological material can accumulate under the fingernail hyporychium of both the victim and/or the suspect and has the potential to provide evidence and intelligence information to the police. The incidence of mixed DNA profiles obtained from fingernail samples Of Couples who co-habit was explored using AmpFlSTR (R) SGM Plus (R) (SGM Plus) (Applied Biosystems (R)). It was predicted that the higher incidence of contact from co-habitation and therefore the greater opportunity for DNA transfer Would result in an increased level of foreign DNA from the partner under the donor's fingernails. Fingernail swabs were taken from all fingers of the left and right hands of 12 couples oil three separate occasions. DNA profiles were compared to both the donor and partner's reference DNA profiles. The number of reportable mixed DNA profiles obtained (17%) was higher than in previous reports. In this Study, the majority of non-donor alleles matched the partner's reference DNA profile, indicating that co-habitation affected non-donor DNA obtained from fingernail swabs. The results demonstrated that as the couples spent increasing amounts of time together, the incidence of mixed DNA profiles increased. The large variation in the incidence of foreign alleles observed within and between Couples suggested that a combination of lifestyle factors were having a significant effect on whether mixed DNA profiles were observed. Further DNA profiling using the AnipFISTR (R) Y-filer (R) (Yfiler) PCR amplification (Applied Biosystems) was carried out to determine whether female fingernail samples that had previously given single donor profiles contained low levels of male DNA that may be detected by targeting the Y-chromosome. It was found that 63% of the samples analysed using Yfiler produced full or partial Y-STR profiles. The Y-STR profiles were compared to partners' profiles and searched through two haplotype databases to determine the evidential value of such samples

Particular genomic and virulence traits associated with preterm infant-derived toxigenic Clostridium perfringens strains.

Clostridium perfringens is an anaerobic toxin-producing bacterium associated with intestinal diseases, particularly in neonatal humans and animals. Infant gut microbiome studies have recently indicated a link between C. perfringens and the preterm infant disease necrotizing enterocolitis (NEC), with specific NEC cases associated with overabundant C. perfringens termed C. perfringens-associated NEC (CPA-NEC). In the present study, we carried out whole-genome sequencing of 272 C. perfringens isolates from 70 infants across 5 hospitals in the United Kingdom. In this retrospective analysis, we performed in-depth genomic analyses (virulence profiling, strain tracking and plasmid analysis) and experimentally characterized pathogenic traits of 31 strains, including 4 from CPA-NEC patients. We found that the gene encoding toxin perfringolysin O, pfoA, was largely deficient in a human-derived hypovirulent lineage, as well as certain colonization factors, in contrast to typical pfoA-encoding virulent lineages. We determined that infant-associated pfoA+ strains caused significantly more cellular damage than pfoA- strains in vitro, and further confirmed this virulence trait in vivo using an oral-challenge C57BL/6 murine model. These findings suggest both the importance of pfoA+ C. perfringens as a gut pathogen in preterm infants and areas for further investigation, including potential intervention and therapeutic strategies

STRmix™ collaborative exercise on DNA mixture interpretation

An intra and inter-laboratory study using the probabilistic genotyping (PG) software STRmix™ is reported. Two complex mixtures from the PROVEDIt set, analysed on an Applied Biosystems™ 3500 Series Genetic Analyzer, were selected. 174 participants responded. For Sample 1 (low template, in the order of 200 rfu for major contributors) five participants described the comparison as inconclusive with respect to the POI or excluded him. Where LRs were assigned, the point estimates ranging from 2 × 10 <sup>4</sup> to 8 × 10 <sup>6</sup> . For Sample 2 (in the order of 2000 rfu for major contributors), LRs ranged from 2 × 10 <sup>28</sup> to 2 × 10 <sup>29</sup> . Where LRs were calculated, the differences between participants can be attributed to (from largest to smallest impact): This study demonstrates a high level of repeatability and reproducibility among the participants. For those results that differed from the mode, the differences in LR were almost always minor or conservative