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Anaplasma (A.) ovis is the most important cause of anaplasmosis in small ruminants. The current study was planned to estimate the molecular prevalence, risk factors, and phylogenetic analysis of A. ovis infection in sheep and goats from different agro-climatic regions of Central and Southern Punjab, Pakistan. A total of 400 jugular blood samples were collected from asymptomatic goats (n = 200) and sheep (n = 200) from the Jhang and Dera Ghazi Khan districts from January 2021 to February, 2023. Two hundred blood samples were collected from each district. Ten union councils (UC) were randomly chosen from each district, and 20 samples were collected from each UC based on the multistage cluster sampling technique. The samples were analyzed with PCR targeting the major surface protein (msp4) gene of A. ovis. The overall molecular prevalence of anaplasmosis was 57.5%. The disease occurrence was higher in Dera Ghazi Khan (61.5%) than in the Jhang district (53.5%). Infection positivity was greater in goats (65.5%) than in sheep (49.5%). Multivariate logistic regression analysis indicated that host species [sheep; Odds Ratio (OR) = 3.212; p = 0.000, Confidence Interval (CI) = 1.968–5.242], age (adult; OR = 2.606; p = 0.003, CI = 1.398–4.858), and acaricide use (never; OR = 13.671; p = 0.000, CI = 6.414–26.283) were significantly higher risk for A. ovis in small ruminants (p< 0.05; OR > 1). The sequencing and phylogenetic analysis of four representative isolates in the current study (Genbank numbers; Goats: OQ302202, OQ302203; Sheep: OQ319592, OQ319593) revealed novel strains of A. ovis with 97–100% similarity from different countries. The msp4-based goat isolates showed greater genetic diversity, while sheep genotypes showed homology with isolates from Italy, Spain, Hungary, Cyprus, Spain, Iran, and China. The current surveillance study will help in devising prevention and control strategies regarding anaplasmosis in small ruminants. However, there is a need for further study on the clinicopathological and vector competence aspects of these genotypes.

Brucellosis is one of the most important worldwide zoonoses of many countries including Egypt. Camel brucellosis has not gained much attention in Egypt yet. This study is focused on the three governorates with the highest camel populations and the largest camel markets in the country to determine the disease seroprevalence and identify the Brucella species in local camel holdings. In total, 381 serum samples were collected from male and female camels from Giza, Aswan, and Al-Bahr Al-Ahmar (the Red Sea) governorates. Samples were serologically examined using the Rose–Bengal plate test (RBPT), indirect ELISA (i-ELISA), competitive ELISA (c-ELISA) and complement fixation test (CFT). Brucella antibodies were detected in 59 (15.5%), 87 (22.8%), 77 (20.2%) and 118 (31.0%) of sera by RBPT, i-ELISA, c-ELISA and CFT, respectively. Using real-time PCR, Brucella DNA was amplified in 32 (8.4%) seropositive samples including Brucella abortus (25/32), Brucella suis (5/32) and Brucella melitensis (2/32), defining a complex epidemiological status. To the best of our knowledge, this is the first study reporting Brucella suis DNA in camel serum. The risk-associated factors including age, sex, breed and geographical distribution were statistically analyzed, showing non-significant association with seroprevalence. The results of this study will raise awareness for camel brucellosis and help develop effective control strategies.

Brucellosis is a highly contagious zoonosis worldwide with economic and public health impacts. The aim of the present study was to identify Brucella (B.) spp. isolated from animal populations located in different districts of Egypt and to determine their antimicrobial resistance. In total, 34-suspected Brucella isolates were recovered from lymph nodes, milk, and fetal abomasal contents of infected cattle, buffaloes, sheep, and goats from nine districts in Egypt. The isolates were identified by microbiological methods and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Differentiation and genotyping were confirmed using multiplex PCR for B. abortus, Brucella melitensis, Brucella ovis, and Brucella suis (AMOS) and Bruce-ladder PCR. Antimicrobial susceptibility testing against clinically used antimicrobial agents (chloramphenicol, ciprofloxacin, erythromycin, gentamicin, imipenem, rifampicin, streptomycin, and tetracycline) was performed using E-Test. The antimicrobial resistance-associated genes and mutations in Brucella isolates were confirmed using molecular tools. In total, 29 Brucella isolates (eight B. abortus biovar 1 and 21 B. melitensis biovar 3) were identified and typed. The resistance of B. melitensis to ciprofloxacin, erythromycin, imipenem, rifampicin, and streptomycin were 76.2%, 19.0%, 76.2%, 66.7%, and 4.8%, respectively. Whereas, 25.0%, 87.5%, 25.0%, and 37.5% of B. abortus were resistant to ciprofloxacin, erythromycin, imipenem, and rifampicin, respectively. Mutations in the rpoB gene associated with rifampicin resistance were identified in all phenotypically resistant isolates. Mutations in gyrA and gyrB genes associated with ciprofloxacin resistance were identified in four phenotypically resistant isolates of B. melitensis. This is the first study highlighting the antimicrobial resistance in Brucella isolated from different animal species in Egypt. Mutations detected in genes associated with antimicrobial resistance unravel the molecular mechanisms of resistance in Brucella isolates from Egypt. The mutations in the rpoB gene in phenotypically resistant B. abortus isolates in this study were reported for the first time in Egypt.

Brucellosis is an infectious disease caused by bacteria of the genus Brucella (B.), affecting both animals and humans, causing severe economic loses and severe illness, respectively. The objective of the present study was to determine the seroprevalence and the risk factors associated with caprine, ovine, and bovine brucellosis in selected districts of Punjab, Pakistan. A total of 1083 blood samples were randomly collected from animals (goats = 440, sheep = 203, cows = 206, and buffaloes = 234). Questionnaires were used to collect data on risk factors associated with brucellosis on the sampling day. All samples were initially screened for anti-Brucella antibodies using the rose bengal plate test (RBPT). The seropositive serum samples were confirmed by a quantitative real-time polymerase chain reaction (PCR) assay for the detection of the Brucella genus- and Brucella species-specific DNA (B. abortus and B. melitensis). Univariant and binary logistic regression were used to identify important risk factors of brucellosis. Anti-Brucella antibodies and DNA were detected in 35 (3.23%) serum samples. Thirty-four (97.1%) DNA samples were confirmed as B. melitensis by qRT-PCR. Abortion history and natural mating were found to be potential risk factors. Brucella melitensis was identified as the causative agent of caprine, ovine, and bovine brucellosis in the selected districts of Punjab, Pakistan. Diseased animals may act as a source of infection for other animals. The elimination of positive seroreactors, development of control strategies for brucellosis, and education programs regarding the control of zoonotic disease are highly needed in developing countries like Pakistan.