diff --git a/modules/hisat2/align/main.nf b/modules/hisat2/align/main.nf
index 8b6eb331..a6f81a6c 100644
--- a/modules/hisat2/align/main.nf
+++ b/modules/hisat2/align/main.nf
@@ -3,10 +3,10 @@ process HISAT2_ALIGN {
     label 'process_high'
 
     // WARN: Version information not provided by tool on CLI. Please update version string below when bumping container versions.
-    conda (params.enable_conda ? "bioconda::hisat2=2.2.0 bioconda::samtools=1.15.1" : null)
+    conda (params.enable_conda ? "bioconda::hisat2=2.2.1 bioconda::samtools=1.15.1" : null)
     container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ?
-        'https://depot.galaxyproject.org/singularity/mulled-v2-a97e90b3b802d1da3d6958e0867610c718cb5eb1:0e773bb207600fcb4d38202226eb20a33c7909b6-0' :
-        'quay.io/biocontainers/mulled-v2-a97e90b3b802d1da3d6958e0867610c718cb5eb1:0e773bb207600fcb4d38202226eb20a33c7909b6-0' }"
+        'https://depot.galaxyproject.org/singularity/mulled-v2-a97e90b3b802d1da3d6958e0867610c718cb5eb1:38aed4501da19db366dc7c8d52d31d94e760cfaf-0' :
+        'quay.io/biocontainers/mulled-v2-a97e90b3b802d1da3d6958e0867610c718cb5eb1:38aed4501da19db366dc7c8d52d31d94e760cfaf-0' }"
 
     input:
     tuple val(meta), path(reads)
@@ -25,7 +25,7 @@ process HISAT2_ALIGN {
     script:
     def args = task.ext.args ?: ''
     def prefix = task.ext.prefix ?: "${meta.id}"
-    def VERSION = '2.2.0' // WARN: Version information not provided by tool on CLI. Please update this string when bumping container versions.
+    def VERSION = '2.2.1' // WARN: Version information not provided by tool on CLI. Please update this string when bumping container versions.
 
     def strandedness = ''
     if (meta.strandedness == 'forward') {
diff --git a/modules/hisat2/build/main.nf b/modules/hisat2/build/main.nf
index 1b36c6b8..fee4064c 100644
--- a/modules/hisat2/build/main.nf
+++ b/modules/hisat2/build/main.nf
@@ -44,7 +44,7 @@ process HISAT2_BUILD {
         log.info "[HISAT2 index build] Less than ${hisat2_build_memory} GB available, so NOT using splice sites and exons to build HISAT2 index."
         log.info "[HISAT2 index build] Use --hisat2_build_memory [small number] to skip this check."
     }
-    def VERSION = '2.2.0' // WARN: Version information not provided by tool on CLI. Please update this string when bumping container versions.
+    def VERSION = '2.2.1' // WARN: Version information not provided by tool on CLI. Please update this string when bumping container versions.
     """
     mkdir hisat2
     $extract_exons
diff --git a/modules/hisat2/extractsplicesites/main.nf b/modules/hisat2/extractsplicesites/main.nf
index d644bcc5..1423f25c 100644
--- a/modules/hisat2/extractsplicesites/main.nf
+++ b/modules/hisat2/extractsplicesites/main.nf
@@ -20,7 +20,7 @@ process HISAT2_EXTRACTSPLICESITES {
 
     script:
     def args = task.ext.args ?: ''
-    def VERSION = '2.2.0' // WARN: Version information not provided by tool on CLI. Please update this string when bumping container versions.
+    def VERSION = '2.2.1' // WARN: Version information not provided by tool on CLI. Please update this string when bumping container versions.
     """
     hisat2_extract_splice_sites.py $gtf > ${gtf.baseName}.splice_sites.txt
     cat <<-END_VERSIONS > versions.yml
diff --git a/tests/modules/hisat2/align/test.yml b/tests/modules/hisat2/align/test.yml
index 54e263bc..a99b2f94 100644
--- a/tests/modules/hisat2/align/test.yml
+++ b/tests/modules/hisat2/align/test.yml
@@ -7,7 +7,6 @@
     - path: output/hisat2/test.hisat2.summary.log
       md5sum: 7b8a9e61b7646da1089b041333c41a87
     - path: output/hisat2/genome.splice_sites.txt
-      md5sum: d41d8cd98f00b204e9800998ecf8427e
     - path: output/hisat2/test.bam
     - path: output/hisat2/hisat2/genome.5.ht2
       md5sum: 91198831aaba993acac1734138c5f173
@@ -35,7 +34,6 @@
     - path: output/hisat2/test.hisat2.summary.log
       md5sum: 9839b31db795958cc4b70711a3414e9c
     - path: output/hisat2/genome.splice_sites.txt
-      md5sum: d41d8cd98f00b204e9800998ecf8427e
     - path: output/hisat2/test.bam
     - path: output/hisat2/hisat2/genome.5.ht2
       md5sum: 91198831aaba993acac1734138c5f173
diff --git a/tests/modules/hisat2/build_test/test.yml b/tests/modules/hisat2/build_test/test.yml
index da5a450c..34c036d0 100644
--- a/tests/modules/hisat2/build_test/test.yml
+++ b/tests/modules/hisat2/build_test/test.yml
@@ -5,7 +5,6 @@
     - hisat2/build
   files:
     - path: output/hisat2/genome.splice_sites.txt
-      md5sum: d41d8cd98f00b204e9800998ecf8427e
     - path: output/hisat2/hisat2/genome.5.ht2
       md5sum: 91198831aaba993acac1734138c5f173
     - path: output/hisat2/hisat2/genome.7.ht2
diff --git a/tests/modules/hisat2/extractsplicesites/test.yml b/tests/modules/hisat2/extractsplicesites/test.yml
index a528199c..2571af94 100644
--- a/tests/modules/hisat2/extractsplicesites/test.yml
+++ b/tests/modules/hisat2/extractsplicesites/test.yml
@@ -5,4 +5,3 @@
     - hisat2/extractsplicesites
   files:
     - path: output/hisat2/genome.splice_sites.txt
-      md5sum: d41d8cd98f00b204e9800998ecf8427e