Active surveillance was conducted by collecting questing ticks from vegetation through a 2-yr survey in Pike County, Pennsylvania. Over a thousand blacklegged ticks (Ixodes scapularis Say) and American dog ticks (Dermacentor variabilis Say) were collected. A single specimen of the following species was collected: lone star tick (Amblyomma americanum L.), rabbit tick (Haemaphysalis leporispalustris Packard), and an Asian longhorned tick (Haemaphysalis longicornis Neumann). This study represents the largest county-wide study in Pennsylvania, surveying 988 questing I. scapularis adult and nymphs. Molecular detection of five distinct tick-borne pathogens was screened through real-time PCR at a single tick resolution. Respectively, the overall 2-yr adult and nymph prevalence were highest with Borrelia burgdorferi (Spirochaetales: Spirochaetacceae) (45.99%, 18.94%), Anaplasma phagocytophilum (Rickettsiales: Anaplasmataceae) (12.29%, 7.95%) where the variant-ha (8.29%, 3.03%) was overall more prevalent than the variant-v1 (2.49%, 4.17%), Babesia microti (Piroplasmida: Babesiidae) (4.97%, 5.30%), Borrelia miyamotoi (Spirochaetales: Spirochaetaceae) (1.38%, 1.89%), and Powassan virus lineage II [POWV]/deer tick virus (DTV) (2.07%, 0.76%). Adult and nymph coinfection prevalence of B. burgdorferi and B. microti (3.03%, 4.97%) and adult coinfection of B. burgdorferi and A. phagocytophilum or A. phagocytophilum and B. microti were significantly higher than the independent infection rate expected naturally. This study highlights the urgency to conduct diverse surveillance studies with large sample sizes to better understand the human risk for tick-borne diseases within small geographical areas.

Babesia microti is a protozoan that infects red blood cells, causing hemolytic anemia and flu-like symptoms in humans. Understanding co-infections is crucial for the better diagnosis, treatment, and management of tick-borne diseases. This study examined the prevalence of Babesia microti co-infection with other prevalent tick-borne pathogens in Pennsylvania. The dataset acquired from the Dr. Jane Huffman Wildlife Genetics Institute included passive surveillance data from Ixodes spp. from 2021 to 2023. Submitted ticks were screened for tick-borne pathogens using species-specific TaqMan qPCR. Of the 793 B. microti-positive ticks pulled for analysis, 65.0% were co-infected with other pathogens (n = 516). Notably, 60.9% of the B. microti-positive ticks were co-infected with Borrelia burgdorferi, 10.2% with Anaplasma phagocytophilum Ap-ha, and 7.5% carried a triple co-infection with B. burgdorferi and A. phagocytophilum Ap-ha. The rates of B. microti infection and its co-infections are on the rise, with patterns observed in Pennsylvania and other regions of the USA. While other studies have collected both nymphal and adult ticks to screen for co-infections in Pennsylvania, our study stood out as a unique contribution to the field by focusing exclusively on B. microti-positive ticks. The continued monitoring of tick-borne co-infections is vital to prevent misdiagnosis and ensure effective treatment regimens.