# Deriving fibroblast cell lines from wing-punch biopsies of Australian eastern bent-winged bats (Miniopterus orianae oceanensis)

## TL;DR

Researchers developed a method to derive fibroblast cell lines from non-lethal wing-punch biopsies of Australian eastern bent-winged bats, improving field sample collection and cell culture techniques.

## Contribution

A novel method for maximizing cell yield from a single biopsy and an alternative transportation method for field samples in bat cell culture.

## Key findings

- Wing-punch biopsies transported in cool PBS and plated as digested cells remained viable.
- Multiple tissue digestions resulted in faster monolayer formation compared to single digestions.
- Non-lethal sampling techniques proved effective for cell culture in an insectivorous bat species.

## Abstract

Cell cultures are a valuable tool for the study of in vitro disease processes, especially where such processes concern wild and/or threatened animal species. However, the collection of adequate samples for cell line preparation can be challenging under field conditions due to logistical challenges and access to equipment. In this paper, we describe the generation of fibroblast cell lines derived from wing-punch biopsies of Australian eastern bent-winged bats (Miniopterus orianae oceanensis), expanding on and modifying existing protocols. Twenty wing-punch biopsies were collected from free-ranging individuals in New South Wales in February 2024 and shipped to the University of Melbourne, Victoria, within 24 hours. To assess the impact of different preservation methods on sample integrity, samples were subjected to two different shipping treatments: Ten were snap-frozen immediately upon collection, and the other ten were placed in cool phosphate-buffered saline (PBS) for transport. To test the effect of different plating treatments, samples were plated as either collagenase-digested cells or explants. Although none of the frozen biopsies or explants showed any growth, all samples transported in cool PBS and plated as digested cells remained viable. While agitation of the samples prior to plating resulted in an initially faster rate of cell growth, cells derived from tissue that had been digested multiple times spread across the plate and formed a monolayer significantly faster than those that had been digested only once. This study confirms the effectiveness of existing cell culture protocols using non-lethal sampling techniques in an Australian insectivorous bat species and shows a novel method of maximizing cell yield from a single biopsy. It also introduces an alternative transportation method, beneficial for field sample collection. These cell cultures are essential tools for future studies on disease susceptibility and pathogen responses in bat species, particularly those belonging to the family Miniopteridae. Additionally, they can be used for biobanking efforts, preserving the genetic material of non-model organisms for broader conservation purposes.