diff --git a/changelogs/fragments/44739-assemble-avoid-extra-newline-Py3.yaml b/changelogs/fragments/44739-assemble-avoid-extra-newline-Py3.yaml
new file mode 100644
index 0000000000..2e86942648
--- /dev/null
+++ b/changelogs/fragments/44739-assemble-avoid-extra-newline-Py3.yaml
@@ -0,0 +1,2 @@
+bugfixes:
+  - assemble - avoid extra newline on Python 3 (https://github.com/ansible/ansible/issues/44739)
diff --git a/lib/ansible/modules/files/assemble.py b/lib/ansible/modules/files/assemble.py
index 04538733e8..37c42021d2 100644
--- a/lib/ansible/modules/files/assemble.py
+++ b/lib/ansible/modules/files/assemble.py
@@ -116,7 +116,7 @@ import re
 import tempfile
 
 from ansible.module_utils.basic import AnsibleModule
-from ansible.module_utils.six import b
+from ansible.module_utils.six import b, indexbytes
 from ansible.module_utils._text import to_native
 
 
@@ -148,7 +148,11 @@ def assemble_from_fragments(src_path, delimiter=None, compiled_regexp=None, igno
                 tmp.write(delimiter)
                 # always make sure there's a newline after the
                 # delimiter, so lines don't run together
-                if delimiter[-1] != b('\n'):
+
+                # byte indexing differs on Python 2 and 3,
+                # use indexbytes for compat
+                # chr(10) == '\n'
+                if indexbytes(delimiter, -1) != 10:
                     tmp.write(b('\n'))
 
         tmp.write(fragment_content)