Improved recovery of ancient DNA from subfossil wood - application to the world's oldest Late Glacial pine forest.

Ancient DNA from historical and subfossil wood has a great potential to provide new insights into the history of tree populations. However, its extraction and analysis have not become routine, mainly because contamination of the wood with modern plant material can complicate the verification of genetic information. Here, we used sapwood tissue from 22 subfossil pines that were growing c. 13 000 yr bp in Zurich, Switzerland. We developed and evaluated protocols to eliminate surface contamination, and we tested ancient DNA authenticity based on plastid DNA metabarcoding and the assessment of post-mortem DNA damage. A novel approach using laser irradiation coupled with bleaching and surface removal was most efficient in eliminating contaminating DNA. DNA metabarcoding confirmed which ancient DNA samples repeatedly amplified pine DNA and were free of exogenous plant taxa. Pine DNA sequences of these samples showed a high degree of cytosine to thymine mismatches, typical of post-mortem damage. Stringent decontamination of wood surfaces combined with DNA metabarcoding and assessment of post-mortem DNA damage allowed us to authenticate ancient DNA retrieved from the oldest Late Glacial pine forest. These techniques can be applied to any subfossil wood and are likely to improve the accessibility of relict wood for genome-scale ancient DNA studies.

Sobrevivência de Campylobacter jejuni em amostras de leite pasteurizado e UHT artificialmente contaminados e mantidas sob refrigeração / Viability of Campylobacter jejuni in the refrigerated samples of artificially contaminated pasteurized and UHT milks

Campylobacter é o agente etiológico mais prevalente em gastroenterites de causa alimentar no mundo. Apesar de o leite cru ser fonte de infecção, pouco se conhece sobre as consequências da recontaminação do leite. A viabilidade de Campylobacter jejuni foi avaliada em leites pasteurizados e UHT mantidos sob refrigeração. Ambos os leites foram divididos em cinco porções de 100 mL, inoculados com 101 UFC.mL-1 de C. jejuni e mantidos de 4 ºC a 7 ºC por 48 horas. Repetiu-se o procedimento, utilizando-se inoculações de 102, 103 e 104 UFC.mL-1. As alíquotas foram analisadas imediatamente após inoculação e depois de 24 e 48 h quanto à viabilidade de C. jejuni. O micro-organismo manteve-se viável em todas as amostras, porém no leite pasteurizado houve redução de 1 ciclo log nas contagens após 24 h e baixas contagens após 48 h. Provavelmente,a redução ocorreu pela presença de microbiota neste leite, que competiu ou inibiu o crescimento de C. jejuni. O leite UHT ofereceu boas condições de sobrevivência em todos os períodos. O consumo de leite contaminado, mesmo em armazenamento refrigerado, pode ser fonte de infecção. O micro-organismo manteve-se mais viável no leite UHT quando comparado ao pasteurizado, provavelmente pela ausência de outros micro-organismos competidores.

Campylobacter is the most prevalent etiologic agent of foodborne gastroenteritis worldwide.Although raw milk is an important source of infection, little is known about the consequencesof milk recontamination. This study aimed at verifying the viability of Campylobacter jejuni inpasteurized and UHT milks stored under refrigeration. Both samples were divided into five portionsof 100 mL, inoculated with 101 CFU.mL-1 of C. jejuni and stored at 4 °C - 7 ºC for 48 h. The procedure was repeated using inoculations of 102, 103 and 104 CFU.mL-1. Aliquots were analyzed immediatelyafter inoculation and after 24 and 48 h to evaluate C. jejuni viability. The microorganism their viabilityin all of samples. In pasteurized milk a reduction of 1 log cycle occurred after 24 h and low countsafter 48 h. Probably, the reduction happened due to the occurrence of microbiota in this milk, whichcaused competition or inhibition of microorganism growth. UHT milk offered good conditions forbacteria survival in all of the periods. The consumption of contaminated milk, even stored under refrigeration might be a source of infection. Microorganism were more viable in UHT milk thanin pasteurized one, probably owing to the absence of competing microorganisms.